A Single-Cell Atlas of the Human Healthy Airways

<b>Rationale:</b> The respiratory tract constitutes an elaborate line of defense that is based on a unique cellular ecosystem.<b>Objectives:</b> We aimed to investigate cell population distributions and transcriptional changes along the airways by using single-cell RNA profiling.<b>Methods:</b> We have explored the cellular heterogeneity of the human airway epithelium in 10 healthy living volunteers by single-cell RNA profiling. A total of 77,969 cells were collected at 35 distinct locations, from the nose to the 12th division of the airway tree.<b>Measurements and Main Results:</b> The resulting atlas is composed of a high percentage of epithelial cells (89.1%) but also immune (6.2%) and stromal (4.7%) cells with distinct cellular proportions in different regions of the airways. It reveals differential gene expression between identical cell types (suprabasal, secretory, and multiciliated cells) from the nose (<i>MUC4</i>, <i>PI3</i>, <i>SIX3</i>) and tracheobronchial (<i>SCGB1A1</i>, <i>TFF3</i>) airways. By contrast, cell-type-specific gene expression is stable across all tracheobronchial samples. Our atlas improves the description of ionocytes, pulmonary neuroendocrine cells, and brush cells and identifies a related population of <i>NREP</i>-positive cells. We also report the association of <i>KRT13</i> with dividing cells that are reminiscent of previously described mouse "hillock" cells and with squamous cells expressing <i>SCEL</i> and <i>SPRR1A/B</i>.<b>Conclusions:</b> Robust characterization of a single-cell cohort in healthy airways establishes a valuable resource for future investigations. The precise description of the continuum existing from the nasal epithelium to successive divisions of the airways and the stable gene expression profile of these regions better defines conditions under which relevant tracheobronchial proxies of human respiratory diseases can be developed.