Fig. 2: Effect of Fgfr2 ligands on purified alveolar epithelial progenitors in culture.

Phase images (left panels) and immunostains (right) of epithelial progenitors purified from tips of e16.5 lungs and cultured in Matrigel for the period indicated with media alone (Control, top panels) or media supplemented every two days with Fgf7 (50 ng/ml, middle panels) or Fgf10 (100 ng/ml) and HSPG (100 ng/ml, lower panels). Note an increase in luminal surface area (dashed lines) and basal extrusion (arrowheads) in cultures with Fgf ligands, and differentiation into intermingled cuboidal AT2 (Sftpc^pos, green) and squamous AT1 cells (RAGE^pos, red) at day 4 (“alveolospheres”). In control cultures lacking FGFs (top row) or treated both with Fgf7 and the Fgfr inhibitor FIIN-1 (bottom panel), cells remained bipotent progenitors throughout, as shown by co-expression of both markers. nAT1, nascent AT1 cell; nAT2, nascent AT2 cell. Bars, 50 µm (left panels and right +Fgf7 panel), 10 µm (right top and +Fgf10 panels), and 10 µm (Fgf7 + FIIN-1 panel). Co-treatment with both Fgf7 and Fgf10 (with HSPG) gave similar results as each treatment alone. Quantification at right gives the percent of each cell type (BP), AT1, AT2) in the culture at day 4 determined by immunostaining (n = 348 cells scored for control, 843 for Fgf7-treatment, and 1365 for Fgf10-treatment in experimental triplicate). ***p = 7.7 × 10⁻²⁸ for Fgf7-treatment and 6.4 × 10⁻²⁶ for Fgf10-treatment (Student’s two-sided t-test), BP abundance of either Fgf treatment condition vs control.