Treatment of cystic fibrosis airway cells with CFTR modulators reverses aberrant mucus properties via hydration.
Treatment of cystic fibrosis airway cells with CFTR modulators reverses aberrant mucus properties via hydration.
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DOI：
10.1183/13993003.00185-2021
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发表时间：
2022-03
期刊：
影响因子：
--
通讯作者：
Ehre C
中科院分区：
文献类型：
--
作者：
Morrison CB;Shaffer KM;Araba KC;Markovetz MR;Wykoff JA;Quinney NL;Hao S;Delion MF;Flen AL;Morton LC;Liao J;Hill DB;Drumm ML;O'Neal WK;Kesimer M;Gentzsch M;Ehre C
囊性纤维化（CF）的特点是肺部黏液积聚。虽然有几种假设与CFTR功能障碍有直接关系（即酸性气道表面液体（ASL） pH，低[HCO3−]，气道脱水），但CF黏液的主要生化改变仍不清楚。我们鉴定了一种新的细胞系（CFTR-KO Calu3细胞）和来自G551D或F508del突变受试者的人支气管上皮细胞（HBE）对Ivacaftor和Elexacaftor-Tezacaftor-Ivacaftor （ETI）的反应。采用短路电流（Isc）、qPCR、ASL pH、western blotting （WB）、光散射/折射法（SEC-MALS）、扫描电镜（SEM）、%固体和颗粒跟踪等光谱分析来确定CFTR功能对粘液性质的影响。Calu3细胞中CFTR功能的丧失导致ASL pH酸化和粘液高浓度（脱水）。CFTR在CF HBE细胞中的调节不影响ASL pH和黏液蛋白mRNA的表达，但降低黏液浓度，放松黏液网络超微结构，改善黏液运输。与调节剂处理的细胞相比，通过使用还原剂去除细胞表面残留的粘液，发现在短暂的根尖清洗后，很大一部分气道粘蛋白仍然附着在naïve CF细胞上。延长水合作用，但不使用NaOH或HCO3−碱化缓冲液，正常化黏液恢复到调节剂处理的细胞水平。这些结果表明，气道脱水，而不是酸性pH和/或低[HCO3−]，是造成CF气道异常粘液特性的原因，CFTR调节主要是恢复正常的粘蛋白缠结。
Cystic fibrosis (CF) is characterized by the accumulation of viscous, adherent mucus in the lungs. While several hypotheses invoke a direct relationship with CFTR dysfunction (i.e., acidic airway surface liquid (ASL) pH, low [HCO3−], airway dehydration), the dominant biochemical alteration of CF mucus remains unknown. We characterized a novel cell line (CFTR-KO Calu3 cells) and the responses of human bronchial epithelial (HBE) cells from subjects with G551D or F508del mutations to Ivacaftor and Elexacaftor-Tezacaftor-Ivacaftor (ETI). A spectrum of assays such as short-circuit currents (Isc), qPCR, ASL pH, western blotting (WB), light scattering/refractometry (SEC-MALS), scanning electron microscopy (SEM), % solids, and particle tracking were performed to determine the impact of CFTR function on mucus properties. Loss of CFTR function in Calu3 cells resulted in ASL pH acidification and mucus hyperconcentration (dehydration). Modulation of CFTR in CF HBE cells did not affect ASL pH or mucin mRNA expression, but decreased mucus concentration, relaxed mucus network ultrastructure, and improved mucus transport. In contrast with modulator-treated cells, a large fraction of airway mucins remained attached to naïve CF cells following short apical washes, as revealed by the use of reducing agents to remove residual mucus from the cell surfaces. Extended hydration, but not buffers alkalized with NaOH or HCO3−, normalized mucus recovery to modulator-treated cell levels. These results indicate that airway dehydration, not acidic pH and/or low [HCO3−], is responsible for abnormal mucus properties in CF airways and CFTR modulation predominantly restores normal mucin entanglement.
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DOI：
10.1126/science.1223012
发表时间：
2012-08-24
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
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期刊：
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发表时间：
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影响因子：
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DOI：
10.1164/rccm.201404-0670oc
发表时间：
2014-08-15
影响因子：
24.7
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