Project Summary The lung possesses a low rate of cellular turnover at homeostasis but contains facultative stem cells that can be activated to regenerate injured tissue. Efficient injury repair in the lung is critical to reestablish gas exchange with the cardiovascular system, a process that is necessary to sustain life. Two key players in the gas exchange process in the distal lung are the type I alveolar epithelial cells and the capillary endothelial cells (ECs), which interface closely with each other to enable transfer of oxygen and carbon dioxide between them. During lung regeneration, distinct subpopulations of capillary ECs may possess cellular plasticity that enables them to act as endothelial progenitors to rebuild the gas exchange machinery. In addition, angiocrine signaling originating from ECs in the distal lung may shape the regeneration of other alveolar cell types. Despite the importance of ECs, however, the extent and function of their heterogeneity within the lung remains incompletely understood. This question is critical to our understanding of the maintenance and repair of functional gas exchange. I have profiled pulmonary EC heterogeneity at single-cell resolution using single-cell RNA sequencing in the adult mouse lung at homeostasis and after acute lung injury and have identified several unique EC populations. These include a subset of ECs that express high levels of signaling molecules and preferentially localize to regions of dense alveolar damage, as well as a population of highly proliferative ECs that arises upon acute lung injury. Each population likely contributes to revascularization of the alveolar space during regeneration. Determining the unique functional role of each population within the alveolus, its preferential response to injury, and how each type of EC interacts with alveolar epithelial cells will facilitate a better understanding of how the lung vasculature is rebuilt and gas exchange is reestablished as the lung regenerates. This mechanistic understanding can then be used to facilitate regenerative medicine approaches that target specific EC subtypes or signaling pathways in the lung. This project will expand my training to include key methods and concepts in lung regeneration and in bioinformatics analysis of single-cell genomic datasets. It will take place under the sponsorship of Dr. Edward Morrisey, a leader in the pulmonary biology field who has defined many key transcription factors and signaling pathways in the lung. This work will be conducted at the University of Pennsylvania, a world-class research institution with collaborative investigators, a rich intellectual environment, and extensive resources for the pursuit of biomedical research focusing on pulmonary biology. Together, the research and training plans proposed herein will facilitate a better understanding of pulmonary endothelial cell heterogeneity and its role in regeneration while preparing me for my future career as an independent...