School of Medicine, Örebro University, University Hospital, Clinical Research Centre, SE-70185, Örebro, Sweden.
Cell Biol Int. 2013 Nov;37(11):1149-56. doi: 10.1002/cbin.10146. Epub 2013 Jul 23.
Ambroxol, a mucokinetic anti-inflammatory drug, has been used for treatment of cystic fibrosis (CF). The respiratory epithelium is covered by the airway surface liquid (ASL), the thickness and composition of which is determined by Cl(-) efflux via the cystic fibrosis transmembrane conductance regulator (CFTR) and Na(+) influx via the epithelial Na(+) channel (ENaC). In cells expressing wt-CFTR, ambroxol increased the Cl(-) conductance, but not the bicarbonate conductance of the CFTR channels. We investigated whether treatment with ambroxol enhances chloride transport and/or CFTR and ENaC expression in CF airway epithelial cells (CFBE) cells. CFBE cells were treated with 100 µM ambroxol for 2, 4 or 8 h. mRNA expression for CFTR and ENaC subunits was analysed by real-time polymerase chain reaction (RT-PCR); protein expression was measured by Western blot. The effect of ambroxol on Cl(-) transport was measured by Cl(-) efflux measurements with a fluorescent chloride probe. Ambroxol significantly stimulated Cl(-) efflux from CFBE cells (a sixfold increase after 8 h treatment), and enhanced the expression of the mRNA of CFTR and α-ENaC, and of the CFTR protein. No significant difference was observed in β-ENaC after exposure to ambroxol, whereas mRNA expression of γ-ENaC was reduced. No significant effects of ambroxol on the ENaC subunits were observed by Western blot. Ambroxol did not significantly affect the intracellular Ca(2+) concentration. Upregulation of CFTR and enhanced Cl(-) efflux after ambroxol treatment should promote transepithelial ion and water transport, which may improve hydration of the mucus, and therefore be beneficial to CF-patients.
氨溴索是一种黏液动力学抗炎药，已被用于治疗囊性纤维化（CF）。呼吸道上皮细胞被气道表面液体（ASL）覆盖，其厚度和成分由氯离子通过囊性纤维化跨膜电导调节因子（CFTR）的外流和钠离子通过上皮钠通道（ENaC）的内流决定。在表达 wt-CFTR 的细胞中，氨溴索增加 CFTR 通道的氯离子电导，但不增加碳酸氢根电导。我们研究了氨溴索治疗是否能增强 CF 气道上皮细胞（CFBE）中氯离子转运和/或 CFTR 和 ENaC 的表达。CFBE 细胞用 100μM 氨溴索处理 2、4 或 8 小时。通过实时聚合酶链反应（RT-PCR）分析 CFTR 和 ENaC 亚基的 mRNA 表达；通过 Western blot 测量蛋白质表达。用荧光氯离子探针测量氯离子流出来测量氨溴索对氯离子转运的影响。氨溴索显著刺激 CFBE 细胞氯离子流出（8 小时处理后增加六倍），并增强 CFTR 和α-ENaC 的 mRNA 表达以及 CFTR 蛋白表达。氨溴索暴露后β-ENaC 无明显变化，而γ-ENaC 的 mRNA 表达减少。Western blot 未观察到氨溴索对 ENaC 亚基的显著影响。氨溴索对细胞内 Ca(2+)浓度没有显著影响。氨溴索治疗后 CFTR 的上调和氯离子流出的增强应促进跨上皮离子和水的转运，从而改善黏液的水合作用，因此对 CF 患者有益。
