Department of Pediatrics, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Pediatr Res. 2011 Nov;70(5):447-52. doi: 10.1203/PDR.0b013e31822f2448.
Cystic fibrosis (CF), a common lethal inherited disorder defined by ion transport abnormalities, chronic infection, and robust inflammation, is the result of mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) protein, a cAMP-activated chloride (Cl-) channel. Macrophages are reported to have impaired activity in CF. Previous studies suggest that Cl- transport is important for macrophage function; therefore, impaired Cl- secretion may underlie CF macrophage dysfunction. To determine whether alterations in Cl- transport exist in CF macrophages, Cl- efflux was measured using N-[ethoxycarbonylmethyl]- 6-methoxy-quinolinium bromide (MQAE), a fluorescent indicator dye. The contribution of CFTR was assessed by calculating Cl- flux in the presence and absence of cftr(inh)-172. The contribution of calcium (Ca(2+))-modulated Cl- pathways was assessed by examining Cl- flux with varied extracellular Ca(2+) concentrations or after treatment with carbachol or thapsigargin, agents that increase intracellular Ca(2+) levels. Our data demonstrate that CFTR contributed to Cl- efflux only in WT macrophages, while Ca(2+)-mediated pathways contributed to Cl- transport in CF and WT macrophages. Furthermore, CF macrophages demonstrated augmented Cl- efflux with increases in extracellular Ca(2+). Taken together, this suggests that Ca(2+)-mediated Cl- pathways are enhanced in CF macrophages compared with WT macrophages.
囊性纤维化（CF）是一种常见的致命遗传性疾病，其特征为离子转运异常、慢性感染和强烈的炎症反应。它是由编码囊性纤维化跨膜电导调节因子（CFTR）蛋白的基因突变引起的，CFTR 蛋白是一种 cAMP 激活的氯离子（Cl-）通道。据报道，巨噬细胞在 CF 中的活性受损。先前的研究表明，Cl-转运对于巨噬细胞功能很重要；因此，Cl-分泌受损可能是 CF 巨噬细胞功能障碍的基础。为了确定 CF 巨噬细胞中是否存在 Cl-转运的改变，使用 N-[乙氧基羰基甲基]-6-甲氧基-喹啉溴化物（MQAE），一种荧光指示剂染料，测量 Cl-外排。通过计算 cftr(inh)-172 存在和不存在时的 Cl-通量来评估 CFTR 的贡献。通过检查不同细胞外 Ca2+浓度或用 carbachol 或 thapsigargin 处理后的 Cl-通量来评估 Ca2+（Ca2+）调节的 Cl-途径的贡献，这些药物可增加细胞内 Ca2+水平。我们的数据表明，CFTR 仅在 WT 巨噬细胞中对 Cl-外排有贡献，而 Ca2+介导的途径在 CF 和 WT 巨噬细胞中均对 Cl-转运有贡献。此外，CF 巨噬细胞在外加 Ca2+增加时表现出增强的 Cl-外排。综上所述，这表明与 WT 巨噬细胞相比，CF 巨噬细胞中 Ca2+ 介导的 Cl-途径增强。
