Rosenblum D A, Volpe M V, Dammann C E, Lo Y S, Thompson J F, Nielsen H C
Division of Newborn Medicine, New England Medical Center, Boston, Massachusetts 02111, USA.
Exp Cell Res. 1998 Feb 25;239(1):69-81. doi: 10.1006/excr.1997.3888.
Epidermal growth factor (EGF) augments late fetal lung maturation by advancing the ontogeny of fetal lung development and by stimulating surfactant synthesis. Previous studies have indicated that fibroblastalveolar epithelial cell communications mediate surfactant synthesis in the fetal lung and EGF acts through such a mechanism. We investigated the hypothesis that is differential activity and expression of the epidermal growth factor receptor (EGF-R) in fetal lung fibroblasts during the canalicular stage of lung development mediates EGF effects. To test this hypothesis, we examined fetal rat lung fibroblasts (FLFs) and type II cells of late gestation (canalicular and saccular stages; 17-22 days) by EGF-R binding techniques, SDS-PAGE, and Western blot analysis. Specific EGF binding increased 181% in day 18 female FLFs, with male FLFs exhibiting a similar increase on day 19. In contrast, specific EGF binding was low in type II cells, did not increase during late gestation, and there were no sex-specific differences. SDS-PAGE and Western blot analysis revealed a predominant 170-kDa EGF-R band in fibroblasts that increased with gestation (peak = 19 days), and was stronger in females. Immunoprecipitation of EGF-treated cells demonstrated the tyrosine kinase activity of the identified receptor. In contrast, type II cells showed minimal signal that did not increase until day 21 of gestation. We also examined whole fetal lung sections by immunohistochemistry to determine cell-specific expression of the EGF-R in vivo. Immunohistochemistry revealed specific EGF-R staining in columnar and cuboidal epithelia of small conducting airways and in mesenchyme of epithelial-mesenchymal borders (including subepithelial mesenchyme). In contrast, alveolar epithelia showed minimal staining, while subalveolar mesenchyme EGF-R staining peaked at day 19 of gestation. We conclude that cell-specific and sex-specific differences in EGF-R binding and EGF-R immunolocalization appears in the fetal lung at a developmental stage that is critical for alveolar epithelial cell differentiation. The results suggest a role for EGF-R activation in late fetal alveolar epithelial cell maturation, which is mediated through mesenchymal-epithelial cell communication.
表皮生长因子（EGF）通过促进胎儿肺发育的个体发生和刺激表面活性剂合成，增强胎儿晚期肺成熟。先前的研究表明，成纤维细胞-肺泡上皮细胞通讯介导胎儿肺中的表面活性剂合成，而EGF通过这种机制发挥作用。我们研究了这样一个假说：在肺发育的小管期，胎儿肺成纤维细胞中表皮生长因子受体（EGF-R）的活性和表达差异介导了EGF的作用。为了验证这一假说，我们通过EGF-R结合技术、SDS-PAGE和蛋白质免疫印迹分析，检测了妊娠晚期（小管期和囊状期；17 - 22天）的胎鼠肺成纤维细胞（FLFs）和II型细胞。在第18天的雌性FLFs中，特异性EGF结合增加了181%，雄性FLFs在第19天表现出类似的增加。相比之下，II型细胞中的特异性EGF结合较低，在妊娠晚期没有增加，并且没有性别特异性差异。SDS-PAGE和蛋白质免疫印迹分析显示，成纤维细胞中主要的170-kDa EGF-R条带随妊娠增加（峰值在第19天），且在雌性中更强。对EGF处理细胞的免疫沉淀证明了所鉴定受体的酪氨酸激酶活性。相比之下，II型细胞显示出最小的信号，直到妊娠第21天才增加。我们还通过免疫组织化学检查了整个胎儿肺切片，以确定体内EGF-R的细胞特异性表达。免疫组织化学显示，在小传导气道的柱状和立方上皮以及上皮-间充质边界的间充质（包括上皮下间充质）中有特异性EGF-R染色。相比之下，肺泡上皮染色最少，而肺泡下间充质EGF-R染色在妊娠第19天达到峰值。我们得出结论，在对肺泡上皮细胞分化至关重要的发育阶段，胎儿肺中EGF-R结合和EGF-R免疫定位存在细胞特异性和性别特异性差异。结果表明，EGF-R激活在胎儿晚期肺泡上皮细胞成熟中起作用，这是通过间充质-上皮细胞通讯介导的。
