Fig. 7.

ISRIB promotes alveolar epithelial differentiation after bleomycin. (A) Schematic of the experiment design. Young adult Sftpc^ERCre × zsGreen mice (3 to 5 mo) received 10 mg of tamoxifen via oral gavage on 2 sequential days 22 d prior to administration of bleomycin (0.025 units, intrathecal). Mice were treated with ISRIB 25 mg/kg intraperitoneally or vehicle 7 d after administration of bleomycin and analyzed 3 d later. (B) Representative flow cytometry data gated on GFP⁺ epithelial cells. (C) The number of GFP⁺PDPN⁺ epithelial cells in vehicle- and ISRIB-treated mice was determined by flow cytometry. Unpaired t test, *P < 0.05. (D) Representative lung sections stained with antibodies against KRT8 and PDPN from mice treated with bleomycin and a single dose of ISRIB or vehicle according to the timeline in A. The arrows indicate GFP⁺KRT8⁺ epithelial cells, and the arrow heads indicate GFP⁺PDPN⁺ AT1 cells. (Scale bar, 100 μm.) (E) Quantification of GFP⁺KRT8⁺ and GFP⁺PDPN⁺ epithelial cells by immunofluorescence. Average cell counts were calculated from at least five nonoverlapping areas per section. Data are shown as mean ± SEM, 4 to 5 mice per group. Mann–Whitney U test. *P < 0.05. (F) Quantification of double positive TUNEL and pro-SPC cells on histological sections after bleomycin. Data are shown as mean ± SEM, 4 to 5 mice per group. Mann–Whitney U test. *P < 0.05.