Department of Physiology, University of Toronto, Toronto, Ontario, Canada.
Am J Respir Cell Mol Biol. 2010 Nov;43(5):599-606. doi: 10.1165/rcmb.2009-0263OC. Epub 2009 Dec 30.
Glucocorticoid (GC)-responsive epithelial-mesenchymal interactions regulate lung development. The GC receptor (GR) mediates GC signaling. Mice lacking GR in all tissues die at birth of respiratory failure. To determine the specific need for epithelial GR in lung development, we bred triple transgenic mice that carry SPC/rtTA, tet-O-Cre, and floxed, but not wild-type, GR genes. When exposed to doxycycline in utero, triple transgenic (GRepi⁻) mice exhibit a Cre-mediated recombination event that inactivates the floxed GR gene in airway epithelial cells. Immunofluorescence confirmed the elimination of GR in Cre-positive airway epithelial cells of late gestation GRepi⁻ mice. Embryonic Day 18.5 pups had a relatively immature appearance with increased lung cellularity and increased pools of glycogen in the epithelium. Postnatal Day 0.5 pups had decreased viability. We used quantitative RT-PCR to demonstrate that specific elimination of epithelial immunoreactive GR in GRepi⁻ mice is associated with reduced mRNA expression for surfactant proteins (SPs) A, B, C, and D; β- and γ-ENaC; T1α; the 10-kD Clara cell protein (CCSP); and aquaporin 5 (AQP5). Western blots confirmed reduced levels of AQP5 protein. No reduction in the levels of the GR transport protein importin (IPO)-13 was observed. Our findings demonstrate a requirement for lung epithelial cell GR in normal lung development. We speculate that impaired epithelial differentiation, leading to decreased SPs, transepithelial Na, and liquid absorption at birth, may contribute to the reduced survival of newborn mice with suppressed lung epithelial GR.
糖皮质激素（GC）反应性上皮-间充质相互作用调节肺发育。GC 受体（GR）介导 GC 信号。所有组织中缺乏 GR 的小鼠在呼吸衰竭的出生时死亡。为了确定上皮 GR 在肺发育中的特定需求，我们培育了三重转基因小鼠，它们携带 SPC/rtTA、tet-O-Cre 和 floxed，但不是野生型，GR 基因。当在子宫内暴露于强力霉素时，三重转基因（GRepi⁻）小鼠表现出 Cre 介导的重组事件，该事件使气道上皮细胞中的 floxed GR 基因失活。免疫荧光证实了晚期妊娠 GRepi⁻小鼠中 Cre 阳性气道上皮细胞中 GR 的消除。胚胎第 18.5 天的幼崽外观相对不成熟，肺泡细胞增多，上皮中糖原池增加。出生后第 0.5 天的幼崽存活能力下降。我们使用定量 RT-PCR 证明，GRepi⁻ 小鼠中上皮免疫反应性 GR 的特异性消除与表面活性剂蛋白（SP）A、B、C 和 D；β-和γ-ENaC；T1α；10-kD 克拉拉细胞蛋白（CCSP）；和水通道蛋白 5（AQP5）的 mRNA 表达减少有关。Western blot 证实 AQP5 蛋白水平降低。未观察到 GR 转运蛋白 importin（IPO）-13 的水平降低。我们的发现表明，正常肺发育需要肺上皮细胞 GR。我们推测，上皮分化受损，导致出生时 SPs、跨上皮 Na 和液体吸收减少，可能导致肺上皮 GR 受抑制的新生小鼠存活率降低。
