471 - The Role of Extracellular Vesicles in Lung Development and Injury

Poster Number: 471
Publication Number: 471.432

Meaghan Ransom, Monroe Carell Jr. Children's Hospital at Vanderbilt, Nashville, TN, United States; Kaitlyn E. Bunn, Vanderbilt University School of Medicine, Nashville, TN, United States; Nicholas M. Negretti, Vanderbilt University Medical Center, Nashville, TN, United States; Christopher S. Jetter, Monroe Carell Jr. Children's Hospital at Vanderbilt, Murfreesboro, TN, United States; John Benjamin, Vanderbilt University Medical Center, Nashville, TN, United States; Dawn Newcomb, Vanderbilt University School of Medicine, Nashville, TN, United States; Heather H. Pua, Vanderbilt University School of Medicine, Nashville, TN, United States; Jennifer Sucre, Vanderbilt University School of Medicine, Nashville, TN, United States

Background: There are currently no available disease-modifying treatments to prevent or treat bronchopulmonary dysplasia (BPD). Extracellular vesicles (EVs) are secreted lipid-enclosed particles that have potential as biomarkers and therapeutic agents in lung disease. While increased EVs have been associated with BPD, little is known about the role of EVs in lung development or after neonatal injury.

Objective: To define the profile of EVs isolated from human tracheal aspirates (TAs) across developmental time, and to investigate changes in the EV secretion profile in mechanical ventilation associated lung injury in premature infants and mice.

Design/Methods: For human studies, tracheal aspirates (TAs) were collected as a part of standard of care from intubated preterm infants, born between 22 and 35 weeks gestation. Samples were analyzed using bead-based flow cytometry with the MACSPlex Exosome Kit. To study injury, adult mice were mechanically ventilated by Flexivent for 20 min. Bronchoalveolar lavage fluid was collected; EVs were analyzed using single-vesicle flow cytometry. Further, our lab recently completed a single-cell transcriptomic atlas of mouse lung development and saccular stage injury (hyperoxia plus LPS), sampling 9 time points from the pseudoglandular through the alveolar stages. We queried the expression data obtained from TAs in this atlas to characterize EV-marker expression.

Results: We collected 49 human samples from 23 patients with gestational ages ranging from 22w1d to 34w6d. 39 samples (80%) from 17 patients (74%) contained detectible EVs. EV tetraspanins CD81 and CD63 were abundant, and lower levels of CD9 were identified. These EVs had high expression of epithelial markers CD326 (EpCAM) and CD133 (Prominin-1). Correlation of marker subsets over time suggests heterogenous vesicle subpopulations within this complex biofluid sample. Comparing ventilated with control mice, we observed a trend toward increased epithelial-derived EVs. Finally, analysis of single-cell RNA sequencing of 17,517 epithelial cells, we found that alveolar type 2 cells have a decrease in CD81 during the saccular stage, and alveolar type 1 cells increase expression of CD81 in the setting of neonatal injury.Conclusion(s): EVs can be detected in tracheal aspirates of preterm infants at birth and specific markers have variable expression with developmental time and in the setting of lung injury. Future work will functionally characterize EVs detected and determine their role in signaling during development and injury, leveraging them as a new therapeutic target in BPD.