Non-Genomic Estrogen Regulation of Ion Transport and Airway Surface Liquid Dynamics in Cystic Fibrosis Bronchial Epithelium

Male cystic fibrosis (CF) patients survive longer than females and lung exacerbations in CF females vary during the estrous cycle. Estrogen has been reported to reduce the height of the airway surface liquid (ASL) in female CF bronchial epithelium. Here we investigated the effect of 17β-estradiol on the airway surface liquid height and ion transport in normal (NuLi-1) and CF (CuFi-1) bronchial epithelial monolayers. Live cell imaging using confocal microscopy revealed that airway surface liquid height was significantly higher in the non-CF cells compared to the CF cells. 17β-estradiol (0.1–10 nM) reduced the airway surface liquid height in non-CF and CF cells after 30 min treatment. Treatment with the nuclear-impeded Estrogen Dendrimer Conjugate mimicked the effect of free estrogen by reducing significantly the airway surface liquid height in CF and non-CF cells. Inhibition of chloride transport or basolateral potassium recycling decreased the airway surface liquid height and 17β-estradiol had no additive effect in the presence of these ion transporter inhibitors. 17β-estradiol decreased bumetanide-sensitive transepithelial short-circuit current in non-CF cells and prevented the forskolin-induced increase in ASL height. 17β-estradiol stimulated an amiloride-sensitive transepithelial current and increased ouabain-sensitive basolateral short-circuit current in CF cells. 17β-estradiol increased PKCδ activity in CF and non-CF cells. These results demonstrate that estrogen dehydrates CF and non-CF ASL, and these responses to 17β-estradiol are non-genomic rather than involving the classical nuclear estrogen receptor pathway. 17β-estradiol acts on the airway surface liquid by inhibiting cAMP-mediated chloride secretion in non-CF cells and increasing sodium absorption via the stimulation of PKCδ, ENaC and the Na+/K+ATPase in CF cells.男性囊性纤维化（CF）患者的生存期长于女性，且CF女性患者的肺部急性加重症状会随发情周期发生变化。已有研究证实，雌激素可降低CF女性支气管上皮的气道表面液体（ASL）高度。本研究探讨了17β-雌二醇对正常（NuLi-1）与CF（CuFi-1）支气管上皮单层细胞的气道表面液体高度及离子转运的影响。采用共聚焦显微镜开展的活细胞成像结果显示，非CF细胞的气道表面液体高度显著高于CF细胞。经0.1~10 nM的17β-雌二醇处理30分钟后，非CF与CF细胞的气道表面液体高度均显著降低。使用核转运受阻的雌激素树状聚合物缀合物（Estrogen Dendrimer Conjugate）进行处理，可模拟游离雌激素的作用，显著降低CF与非CF细胞的气道表面液体高度。抑制氯离子转运或基底侧钾离子循环可降低气道表面液体高度，且在此类离子转运抑制剂存在的条件下，17β-雌二醇未表现出附加效应。17β-雌二醇可降低非CF细胞中布美他尼敏感的跨上皮短路电流，并阻断福司柯林诱导的ASL高度升高。在CF细胞中，17β-雌二醇可激活阿米洛利敏感的跨上皮电流，并提升哇巴因敏感的基底侧短路电流。17β-雌二醇可增强CF与非CF细胞中的蛋白激酶Cδ（PKCδ）活性。上述实验结果证实，雌激素可使CF与非CF细胞的ASL脱水，且细胞对17β-雌二醇的此类应答属于非基因组效应，而非依赖经典核雌激素受体通路。17β-雌二醇通过以下方式作用于气道表面液体：在非CF细胞中抑制cAMP介导的氯离子分泌，在CF细胞中通过激活PKCδ、上皮钠通道（ENaC）与钠钾ATP酶（Na+/K+ATPase）来提升钠吸收效率。