Extended Data Fig. 3: Epithelial clusters in the normal human lung proximal airways lack the distinct secretory subtypes found in distal ariways.

A) Heatmap of cluster defining genes within the proximal human epithelium. B) Selected gene expression compared across all proximal epithelial clusters shown as dot plot. C) RNAscope analysis for SCGB1A1 and SCGB3A2 transcripts reveals a similar distribution to protein expression in distal airways; A distal airway transitioning from terminal to respiratory bronchiole is shown, highlighting the preserved gradient of SCGB1A1 expressing cells in larger airways to SCGB3A2 expressing cells in more distal airways. Scale bar = 500 µm. N = 3. D, E) Additional airway structure showing intermixing of SCGB1A1 and SCGB3A2 positive populations. Yellow arrows point to double positive cells along gradient. While with the scRNA analysis there is SCGB1A1 expression in SCGB3A2 positive cells, double positive cells were rarely observed in RNAscope or protein IHC, likely due to a threshold of the technique. Yellow arrows indicate rare double positive cells in a distal human respiratory airway. Red arrows indicate SCGB3A2 single positive cells. Scale bars = 50 µm. N = 3. F, G) IHC of SCGB1A1 and SCGB3A2 expression in large and distal airways from representative individual patients (n = 6). Scale bar = 100 µm.