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. 2022 Dec 8;207(8):1042–1054. doi: 10.1164/rccm.202207-1332OC

Figure 5.

Figure 5.

TanFe (transcellular activator of nuclear FOXF1 expression) increases lung angiogenesis and prevents mortality in Foxf1+/− (Forkhead Box F1) mice. (A) Schematic shows a breeding strategy and TanFe treatments of pregnant mice on Embryonic Days E11.5, E13.5, and E15.5 (10 mg/kg body weight). (B) TanFe increases FOXF1 protein amounts in lungs of Foxf1+/− embryos. Immunoblots show the concentrations of FOXF1, FLK1, PECAM-1, FAAP100, and β-Actin in lung extracts from E18.5 embryos treated with either TanFe or vehicle. Protein lysates from three WT lungs were pooled together before the analysis. Foxf1+/− lungs (Het) were analyzed individually (left panels). Quantification of two independent Western blots shows that TanFe does not change FAAP100 protein concentrations but increases FOXF1, FLK1, and PECAM1 in Foxf1+/− embryos collected at E18.5 to P1 (n = 3–6 embryos in each group) (right graphs). (C) TanFe increases capillary density in Foxf1+/− lungs. Images show endomucin staining (green) of E18.5 lungs after TanFe treatment. DAPI (blue) was used to stain cell nuclei. Magnification is ×400. Quantification of endomucin staining was performed using ImageJ software in 10 random images from three mouse lungs in each group. (D) TanFe increases survival of Foxf1+/− mice after birth. Mice were treated with TanFe (10 mg/kg body weight) or vehicle on Embryonic Days E11.5, E13.5, and E15.5. Survival rates of vehicle-treated Foxf1+/− mice (n = 175) and TanFe-treated Foxf1+/− mice (n = 49) were determined at Postnatal Day 30 (P30). (E) TanFe improves alveolarization in Foxf1+/− mice after birth. Hematoxylin and eosin staining shows that TanFe decreases alveolar simplification in Foxf1+/− mice at P35 (n = 7–13 in each group). Magnification is ×100. *P < 0.05, **P < 0.01, and ***P < 0.001. n.s. = not significant; WT = wild-type.