Generation of mature lung alveolar epithelial cells from human pluripotent stem cells

We perform time-series global transcriptomic profiling of a directed differentiation protocol for generating alveolar epithelial type II cells (AEC2s) from pluripotent stem cells (PSCs). We analyzed 3 different timepoints in the RUES2 differentiation: 1) Day 0 undifferentiated cells, 2) Day 15 lung progenitors highly enriched in NKX2-1+ cells by CD47hi/CD26lo sorting, and 3) the outgrowth of these purified progenitors in 3D culture sorted again on Day 35 based on SFTPCtdTomato+ and SFTPCtdTomato- gating. For comparison to primary cells, we simultaneously sequenced RNA from purified primary fetal (21 week gestation) distal alveolar epithelial progenitors and purified adult human (HTII-280 sorted) AEC2s. In order to evaluate the effect of cell culture on primary fetal alveolar cells, parallel aliquots of the fetal cells were also exposed to 4 days of culture in DCI media. We find that AEC2 maturation involves downregulation of Wnt signaling activity, and that the highest differentially expressed transcripts in iPSC-derived AEC2s encode genes associated with lamellar body and surfactant biogenesis.