Measurements of spontaneous CFTR-mediated ion transport without acute channel activation in airway epithelial cultures after modulator exposure.
Measurements of spontaneous CFTR-mediated ion transport without acute channel activation in airway epithelial cultures after modulator exposure.
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DOI：
10.1038/s41598-021-02044-1
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发表时间：
2021-11-19
影响因子：
4.6
通讯作者：
Bratcher PE
中科院分区：
文献类型：
--
作者：
Nick HJ;Zeitlin PL;Yadav S;Bratcher PE
CFTR功能的体外定量通常通过急性刺激然后抑制通过CFTR的离子转运并测量所产生的跨上皮电压（Vte）和电流（ISC）的变化来进行。虽然该技术适用于测量CFTR的最大功能能力，但它可能无法提供体内CFTR活性的准确估计。为了测试CFTR介导的离子转运是否可以在不存在急性CFTR刺激的情况下测量，在Ussing室中用阿米洛利处理，然后用CFTR（inh）-172处理，而没有CFTR的急性激活，分析初级气道上皮。非CF上皮细胞在暴露于CFTR（inh）-172和不存在毛喉素/IBMX（F/I）后显示Vte和ISC降低;这种降低被解释为这些上皮细胞中存在的自发CFTR活性的量度。在F508 del/F508 del CFTR上皮细胞中，用VX-809/VX-770处理后，F/I诱导的Vte和ISC变化增加约4倍，而VX-809/VX-770处理后自发CFTR活性的幅度仅增加约1.6倍。观察到其他CF上皮细胞对调节剂治疗的反应存在方法依赖性差异。这些结果用作在不存在急性CFTR活化的情况下体外分析CFTR调节剂应答的概念证明。未来的研究将确定这种方法在开发新型CFTR调节剂疗法中的有用性。
Quantitation of CFTR function in vitro is commonly performed by acutely stimulating then inhibiting ion transport through CFTR and measuring the resulting changes in transepithelial voltage (Vte) and current (ISC). While this technique is suitable for measuring the maximum functional capacity of CFTR, it may not provide an accurate estimate of in vivo CFTR activity. To test if CFTR-mediated ion transport could be measured in the absence of acute CFTR stimulation, primary airway epithelia were analyzed in an Ussing chamber with treatment of amiloride followed by CFTR(inh)-172 without acute activation of CFTR. Non-CF epithelia demonstrated a decrease in Vte and ISC following exposure to CFTR(inh)-172 and in the absence of forskolin/IBMX (F/I); this decrease is interpreted as a measure of spontaneous CFTR activity present in these epithelia. In F508del/F508del CFTR epithelia, F/I-induced changes in Vte and ISC were ~ fourfold increased after treatment with VX-809/VX-770, while the magnitude of spontaneous CFTR activities were only ~ 1.6-fold increased after VX-809/VX-770 treatment. Method-dependent discrepancies in the responses of other CF epithelia to modulator treatments were observed. These results serve as a proof of concept for the analysis of CFTR modulator responses in vitro in the absence of acute CFTR activation. Future studies will determine the usefulness of this approach in the development of novel CFTR modulator therapies.
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期刊：
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影响因子：
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