Prenatal FGFR2 Signaling via PI3K/AKT Specifies the PDGFRA⁺ Myofibroblast

It is well known that Fibroblast Growth Factor Receptor 2 (FGFR2) signaling is critical for proper lung development. Recent studies demonstrate that epithelial FGFR2 signaling during the saccular phase of lung development (sacculation) regulates AT1 and AT2 cell differentiation. During sacculation, Platelet-Derived Growth Factor Receptor Alpha-positive (PDGFRA+) lung fibroblasts exist as three functional subtypes: contractile myofibroblasts, ECM-producing matrix fibroblasts, and lipofibroblasts. All three subtypes are required during alveolarization to establish a niche that supports alveolar type-2 epithelial (AT2) self-renewal and alveolar type-1 epithelial (AT1) differentiation. FGFR2 signaling directs myofibroblast differentiation in PDGFRA+ fibroblasts during alveolar reseptation after pneumonectomy. However, it remains unknown if FGFR2 signaling regulates PDGFRA+ myo-, matrix-, or lipofibroblast differentiation during sacculation. In this study, FGFR2 signaling was inhibited by temporal expression of a secreted dominant-negative FGFR2b (dnFGFR2) by AT2 cells from E16.5 to E18.5. Fibroblast and epithelial differentiation were analyzed at E18.5, PN7, and PN21. At all time points, the number of myofibroblasts was reduced and the number lipo/matrix fibroblasts was increased. AT2 cells are increased and AT1 cells are reduced postnatally, but not at E18.5. Similarly, in organoids made with PDGFRA+ fibroblasts from dnFGFR2 lungs, increased AT2 cells and reduced AT1 cells were observed. In vitro treatment of primary wild-type E16.5 adherent saccular lung fibroblasts with recombinant dnFGFR2b/c resulted in reduced myofibroblast contraction. Treatment with PI3K/AKT activator 740 Y-P rescued the lack of myofibroblast differentiation caused by dnFGFR2b/2c. Moreover, treatment with PI3K/AKT activator 740 Y-P rescued myofibroblast differentiation in E18.5 fibroblasts isolated from dnFGFR2 lungs.