Decreased Expression Levels of Leukemia Inhibitory Factor and Its Receptor During Airway Branching Morphogenesis in Nitrofen-Induced Hypoplastic Lungs

Abstract

Purpose: Pulmonary hypoplasia (PH) remains a major therapeutic challenge associated with congenital diaphragmatic hernia (CDH). Leukemia inhibitory factor (LIF) and its receptor (LIFR) play an essential role in airway branching morphogenesis in developing fetal lungs. With a significant increase just before birth, high LIF expression level has been demonstrated in the pulmonary epithelium, whereas LIFR is mainly expressed in the surrounding mesenchyme. Furthermore, LIFdeficient fetuses exhibit PH with defective airway tissue. We hypothesized that pulmonary LIF and LIFR expression levels are decreased during lung branching morphogenesis in nitrofen-induced PH. Methods: Timed-pregnant rats received either nitrofen or a vehicle on gestational day 9 (D9). Fetuses were harvested on D15, D18 and D21, and dissected lungs were divided into control and nitrofen-exposed groups (n = 12 per time point and group). The pulmonary expression levels of LIF and LIFR were analyzed by quantitative real-time polymerase chain reaction. Immunohistochemical staining of LIF and LIFR was performed to evaluate protein expression and localization in branching airway tissue. Results: Relative mRNA expression levels of LIF and LIFR were significantly reduced in the lungs of nitrofen-exposed fetuses on D15 (0.21 ± 0.13 vs 0.49 ± 0.19; p < 0.05 and 0.24 ± 0.13 vs. 0.36 ± 0.12; p < 0.05), D18 (0.11 ± 0.07 vs. 0.49 ± 0.41; p < 0.05 and 0.10 ± 0.03 vs. 0.16 ± 0.03; p < 0.05) and D21 (0.13 ± 0.04 vs. 0.27 ± 0.05; p < 0.05 and 0.18 ± 0.03 vs. 0.34 ± 0.12; p < 0.05) compared with controls. LIF immunoreactivity was markedly diminished in the distal airway epithelium, whereas LIFR expression level was decreased in mesenchymal cells surrounding terminal bronchioles and alveoli on D15, D18 and D21 compared with controls. Conclusion: Decreased pulmonary LIF and LIFR expression levels may disrupt epithelialmesenchymal interactions during lung branching morphogenesis and cause PH in the nitrofen-induced CDH model.