Effects of dexamethasone, growth factors, and tracheal ligation on the development of nitrofen-exposed hypoplastic murine fetal lungs in organ culture

Abstract

Background/Purpose : The addition of growth factors EGF (epidermal growth factor) plus TGFβ 1 (transforming growth factor β 1; E + T) or dexamethasone (DEX) to normal murine fetal lungs in culture enhances lung development. In addition, ligation of the airway in lungs in organ culture, enhances lung development. Nitrofen (2,4-dichlorophenyl- p-nitrophenylether) administration to pregnant mice results in pulmonary hypoplasia in the offspring with many similarities to human hypoplastic lung conditions. This study investigates the effects of growth factors, dexamethasone, and airway ligation on the development of hypoplastic fetal murine lungs in whole-organ culture. We hypothesized that E+T, DEX, or airway ligation will enhance the development and maturation of hypoplastic murine fetal lungs in vitro. Methods : Time-dated pregnant CD-1 mice were given nitrofen, 25 mg, intragastrically at gestational day (Gd) 8. The dams were killed on Gd 14, and the fetuses were removed. The hypoplastic fetal lungs were excised, and the tracheae were transected. The lungs were cultured in serum-free BGJb media in the presence or absence of E+T (10 ng/mL + 2 ng/mL, respectively) or DEX (10 nmol/L). Some lungs were cultured for 7 days with the tracheae ligated. Results : Gross morphology under a dissecting stereomicroscope showed that the lungs were larger after E+T, DEX, or tracheal ligation. Histologically, the untreated lungs had progressed from the pseudoglandular stage to a canalicular-like stage with poorly differentiated airways. The E+T-treated lungs had better developed airway branching and small acini; however, thick mesenchyme persisted. The ligated lungs had well-developed airway branching and acinar structures. After DEX treatment the lungs were most developed with very well defined airway branching and expanded acinar structures; however, there was no secondary septation. Ultrastructurally, the hypoplastic lungs at Gd 14 and after 7 days in culture had no glycogen in their epithelial cells, no defined acinar formation, and had damaged mitochondria. The E+T-treated or tracheally ligated lungs had abundant type II cells, secreted lamellar bodies (LBs), and showed infrequent tubular myelin. Mitochondrial damage was noted in these lungs as in the untreated lungs. DEX-treated hypoplastic lungs showed large acini. The acinar walls were thick; however, they had type II cells with abundant LBs and intact mitochondria. The airways were noted to have differentiated cell types. Surfactant secretions in acinar spaces showed tubular myelin structures. Conclusions : E+T, tracheal ligation, or DEX accelerates lung development and maturation of hypoplastic fetal murine lungs compared with untreated controls. DEX had a greater effect with special reference to repair of mitochondrial damage. DEX not only accelerated lung development, but it may have reversed some of the effects nitrofen.