Dankó Tamás, Hargitai Dóra, Pataki Agnes, Hakim Hasif, Molnár Miklós, Zsembery Akos
Institute of Human Physiology and Clinical Experimental Research, Semmelweis University, Budapest, Hungary.
Cell Physiol Biochem. 2011;27(3-4):401-10. doi: 10.1159/000327967. Epub 2011 Apr 1.
The pH of the airway surface liquid (ASL) plays a pivotal role in maintaining the proper function of the respiratory epithelium. In patients with cystic fibrosis (CF) acidic ASL has been observed. Thus, alkalinization of ASL itself might be beneficial in CF. The aim of this study was to investigate the role of extracellular pH (pH(o)) on the alternative Ca(2+)-activated Cl(-) channels (CaCCs) in CF airway epithelial cells.
The Ca(2+) and viability of CF airway epithelial cells (IB3-1) were assessed using Fluo-3/AM and YO-PRO-1 fluorescent dyes, respectively. Ion currents were detected in whole-cell configuration using the patch clamp technique.
Extracellular alkalinization (pH(o) 8.2) stimulated Ca(2+) entry and inward currents in low Na(+) containing medium. The inward currents were blocked by the removal of extracellular Ca(2+), chelating cytosolic Ca(2+), as well as by the application of niflumic acid and DIDS. While Zn(2+) promoted sustained Ca(2+) entry in pH(o)-dependent manner, it inhibited the anion conductance. The low external Na(+) concentrations and alkaline pH(o) were well tolerated by the cells.
Stimulation of CaCCs could be achieved by alkalinization of the extracellular environment in CF airway epithelial cells. Zn(2+) directly blocked, however indirectly enhanced the activity of Cl(-) conductance.
气道表面液体（ASL）的pH值在维持呼吸道上皮的正常功能中起着关键作用。在囊性纤维化（CF）患者中已观察到酸性ASL。因此，ASL本身的碱化可能对CF有益。本研究的目的是探讨细胞外pH值（pH(o)）对CF气道上皮细胞中替代性钙激活氯通道（CaCCs）的作用。
分别使用Fluo-3/AM和YO-PRO-1荧光染料评估CF气道上皮细胞（IB3-1）的Ca(2+)和活力。使用膜片钳技术在全细胞模式下检测离子电流。
在含低钠的培养基中，细胞外碱化（pH(o) 8.2）刺激了Ca(2+)内流和内向电流。去除细胞外Ca(2+)、螯合胞质Ca(2+)以及应用尼氟酸和DIDS均可阻断内向电流。虽然Zn(2+)以pH(o)依赖的方式促进持续的Ca(2+)内流，但它抑制了阴离子电导。细胞对低的外部Na(+)浓度和碱性pH(o)具有良好的耐受性。
通过CF气道上皮细胞外环境的碱化可实现对CaCCs的刺激。然而，Zn(2+)直接阻断但间接增强了Cl(-)电导的活性。
