Murine nitrofen-induced pulmonary hypoplasia does not involve induction of TGF-beta signaling.

BACKGROUND/PURPOSE: In the murine nitrofen-induced model of congenital diaphragmatic hernia (CDH), the lungs are primarily hypoplastic and immature even before diaphragmatic closure. Because excess transforming growth factor-beta (TGF-beta) signaling induces pulmonary hypoplasia, the authors hypothesized that primary hypoplasia after nitrofen exposure may be caused by aberrant signaling by the TGF-beta pathway. Therefore, abrogation of TGF-beta signaling might rescue the hypoplasia. METHODS: The authors performed intratracheal microinjections of a recombinant adenoviral vector encoding a dominant-negative TGF-beta type II receptor (AdIIR-DN) in nitrofen-exposed and control E12 mouse lungs, which then were cultured for 4 days in serumless chemically defined media. The mRNA expression of Smad2, 3, 4, and 7 in nitrofen-exposed and control E12 lungs after 4 days in culture were compared. RESULTS: ADIIR-DN increased terminal branching in control lungs by 28% compared with lungs injected with control virus (61.8 +/- 4.6 v. 48.4 +/- 4.7, P =.004). However, there was no difference between nitrofen-exposed lungs injected with ADIIR-DN and those injected with control virus. Compared with control lungs, Smad mRNA expression was decreased markedly in nitrofen-exposed lungs: Smad2 (40%, P =.16), Smad3 (29%, P =.02), Smad4 (25%, P =.07), and Smad7 (36%, P =.04). CONCLUSIONS: Because abrogation of TGF-beta signaling does not rescue the hypoplasia seen in the nitrofen model, and Smad expression is decreased in nitrofen-exposed lungs, the TGF-beta pathway does not appear to play a role in nitrofen-induced pulmonary hypoplasia.

Nitrofen inhibition of pulmonary growth and development occurs in the early embryonic mouse.

BACKGROUND/PURPOSE: It was believed previously that pulmonary hypoplasia in congenital diaphragmatic hernia (CDH) was a consequence of the herniation of abdominal viscera into the chest. Using the murine nitrofen-induced model of CDH, the authors evaluated lung growth and development before diaphragm closure or herniation. METHODS: The authors examined nitrofen-exposed early embryonic lungs on embryonic day 12 (E12). Branching morphogenesis was quantified before and after 4 days in culture in serumless chemically defined media and compared with age-matched control lungs. The mRNA expression of proliferative and developmental markers in cultured lungs was then determined. RESULTS: Nitrofen-exposed lungs had 30% fewer total terminal branches than age-matched controls (9.3 +/- 1.9 nitrofen v 13.7 +/- 2.6 control; P <.001). Hypoplasia also was more profound in the left than the right lung. These effects persisted after culturing the lungs for 4 days in serumless chemically-defined media (31.7 +/- 6.8 nitrofen v 42.9 +/- 8.4 control, P <.001). Furthermore, the mRNA expression of proliferative and developmental markers was decreased in nitrofen-exposed E12 lungs cultured for 4 days (as a percentage of age-matched controls): cyclin A (69.28%; P =.04), Nkx2.1 (44.4%, 0.04), SP-A (24.1%; P =.008), SP-B (23.4%; P =.05), SP-C (20%; P =.06), and CC-10 (13.8%; P =.04). CONCLUSION: Nitrofen induces primary pulmonary hypoplasia and immaturity in the early embryonic mouse, and this effect persists in culture.