Single-Cell Profiling of Premature Neonate Airways Reveals a Continuum of Myeloid Differentiation.

Single-cell genomic technologies hold great potential to advance our understanding of lung development and disease. A major limitation lies in accessing intact cells from primary lung tissues for profiling human airway health. Sampling methods such as endotracheal aspiration that are compatible with clinical interventions could enable longitudinal studies, the enrollment of large cohorts, and the development of novel diagnostics. To explore single-cell RNA sequencing profiling of the cell types present at birth in the airway lumen of extremely premature neonates (<28 wk gestation), we isolated cells from endotracheal aspirates collected from intubated neonates within the first hour after birth. We generated data on 10 subjects, providing a rich view of airway luminal biology at a critical developmental period. Our results show that cells present in the airways of premature neonates primarily represent a continuum of myeloid differentiation, including fetal monocytes (25% of total), intermediate myeloid populations (48%), and macrophages (2.6%). Applying trajectory analysis to the myeloid populations, we identified two trajectories consistent with the developmental stages of interstitial and alveolar macrophages, as well as a third trajectory presenting an alternative pathway bridging the distinct macrophage precursors. The three trajectories share many dynamic genes (