Delpiano Livia, Thomas Joseph J, Yates Annabel R, Rice Sarah J, Gray Michael A, Saint-Criq Vinciane
Epithelial Research Group, Institute for Cell and Molecular Biosciences, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, United Kingdom.
Skeletal Research Group, Institute of Genetic Medicine, International Centre for Life, Newcastle University, Newcastle upon Tyne, United Kingdom.
Front Pharmacol. 2018 Dec 11;9:1462. doi: 10.3389/fphar.2018.01462. eCollection 2018.
Respiratory failure, driven by airways mucus obstruction, chronic inflammation and bacterial infections, is the main cause of mortality and morbidity in people with cystic fibrosis (CF) due to defects in the Cl and transport activity of the CF Transmembrane conductance Regulator (CFTR). Most recent pre-clinical and clinical studies have focused on restoring CFTR function by enhancing its trafficking or transport activity and show promising results. However, there are a significant number of patients that will not benefit from these CFTR-targeted therapies and it is therefore important to identify new non-CFTR targets that will restore lung function, by-passing CFTR dysfunction. The H/K-ATPase, ATP12A, has recently been identified as a potential novel target for CF therapies, since its acute inhibition by ouabain was shown to help restore mucus viscosity, mucociliary transport, and antimicrobial activity using CF airway models, and this effect was linked to an increase in the pH of the airway surface liquid (ASL). Here, we have evaluated the potential therapeutic use of ouabain by investigating the effect of chronically treating fully differentiated CF primary human airway epithelial cells (hAECs) with ouabain, under thin film conditions, resembling the situation. Our results show that although chronic treatment increased ASL pH, this correlated with a deleterious effect on epithelial integrity as assessed by LDH release, transepithelial electrical resistance, fluorescein flux, and ion transport. Since ATP12A shares approximately 65% identity with the gastric H/K-ATPase (ATP4A), we investigated the potential of using clinically approved ATP4A proton pump inhibitors (PPIs) for their ability to restore ASL pH in CF hAECs. We show that, despite not expressing ATP4A transcripts, acute exposure to the PPI esomeprezole, produced changes in intracellular pH that were consistent with the inhibition of H secretion, but this response was independent of ATP12A. More importantly, chronic exposure of CF hAECs to esomeprazole alkalinized the ASL without disrupting the epithelial barrier integrity, but this increase in ASL pH was consistent with a decrease in mRNA expression of . We conclude that PPIs may offer a new approach to restore ASL pH in CF airways, which is independent of CFTR.
由气道黏液阻塞、慢性炎症和细菌感染导致的呼吸衰竭，是囊性纤维化（CF）患者死亡和发病的主要原因，这是由于CF跨膜电导调节因子（CFTR）的Cl和转运活性存在缺陷。最近的临床前和临床研究集中在通过增强CFTR的转运或运输活性来恢复其功能，并显示出有希望的结果。然而，有相当数量的患者无法从这些针对CFTR的治疗中获益，因此确定新的非CFTR靶点以绕过CFTR功能障碍来恢复肺功能很重要。H/K-ATP酶ATP12A最近被确定为CF治疗的潜在新靶点，因为使用CF气道模型显示哇巴因对其急性抑制有助于恢复黏液黏度、黏液纤毛运输和抗菌活性，且这种作用与气道表面液体（ASL）pH值的升高有关。在此，我们通过研究在类似生理情况的薄膜条件下用哇巴因长期处理完全分化的CF原代人气道上皮细胞（hAECs）的效果，评估了哇巴因的潜在治疗用途。我们的结果表明，尽管长期处理会增加ASL的pH值，但这与通过乳酸脱氢酶释放、跨上皮电阻、荧光素通量和离子转运评估的对上皮完整性的有害影响相关。由于ATP12A与胃H/K-ATP酶（ATP4A）约有65%的同源性，我们研究了使用临床批准的ATP4A质子泵抑制剂（PPIs）恢复CF hAECs中ASL pH值的潜力。我们表明，尽管不表达ATP4A转录本，但急性暴露于PPI埃索美拉唑会产生与H分泌抑制一致的细胞内pH值变化，但这种反应与ATP12A无关。更重要的是，CF hAECs长期暴露于埃索美拉唑会使ASL碱化而不破坏上皮屏障完整性，但ASL pH值的这种升高与某种物质的mRNA表达降低一致。我们得出结论，PPIs可能提供一种独立于CFTR恢复CF气道中ASL pH值的新方法。
