A single-cell atlas of human fetal lung development between 14 and 19 weeks of gestation
Rationale. Human lung development has been mainly described in morphologic studies and the potential underlying molecular mechanisms were extrapolated from animal models. Therefore, there is a need to gather knowledge from native human lung tissue. In this study we describe changes at a single-cell level in human fetal lungs during the pseudoglandular stage. Methods. We report the cellular composition, cell trajectories and cell-to-cell communication in developing human lungs with single-nuclei RNA sequencing (snRNA-seq) on 23,251 nuclei isolated from nine human fetuses with gestational ages between 14 to 19 weeks of gestation.  Results. We identified nine main lung cell populations and 21 sub-populations, including a rare pulmonary neuroendocrine cells population. For each cell type, marker genes are reported, and selected marker genes are used for spatial validation with fluorescent RNA in situ hybridization. Enrichment and changes in cell state analysis across the different gestational ages provide insight into molecular mechanisms and signaling pathways within individual cell clusters according to gestational age. Lastly, ligand-receptor analysis highlights determinants of cell-to-cell communication among the different cell types through the pseudoglandular stage, including general developmental pathways (NOTCH and TGFB), as well as more specific pathways involved in vasculogenesis, neurogenesis, and immune system regulation.  Conclusion. These findings provide a clinically relevant background for research hypotheses generation in projects studying normal or impaired lung development and help to develop and validate surrogate models to study human lung development, such as human lung organoids. Overall design: Expression profiling by high throughput sequencing on human fetla lung tissue from 14 to 19 weeks of gestation