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Phospholipase C controls chloride-dependent short-circuit current in human bronchial epithelial cells
American Journal of Physiology-Lung Cellular and Molecular Physiology ( IF 3.6 ) Pub Date : 2020-11-25 , DOI: 10.1152/ajplung.00437.2019
Chloé Grebert 1 , Frederic Becq 2 , Clarisse Vandebrouck 1
Affiliation  

Chloride secretion by airway epithelial cells is primordial for water and ion homeostasis and airways surface prevention of infections. This secretion is impaired in several human diseases, including cystic fibrosis, a genetic pathology due to CFTR gene mutations leading to chloride channel defects. A potential therapeutic approach is aiming at increasing chloride secretion either by correcting the mutated CFTR itself or by stimulating non-CFTR chloride channels at the plasma membrane. Here we studied the role of phospholipase C in regulating the transepithelial chloride secretion in human airway epithelial 16HBE14o- and CFBE cells over expressing WT‑ or F508del‑CFTR. Western blot analysis shows expression of the three endogenous PLC isoforms PLCd1, PLCg1 and PLCb3 in 16HBE14o‑ cells. In 16HBE14o‑ cells we performed Ussing chamber experiments after silencing each of these PLC isoforms or using the PLC inhibitor U73122 or its inactive analogue U73343. Our results show the involvement of PLCb3 and PLCg1 in CFTR-dependent short-circuit current activated by forskolin, but not PLCd1. In CFBE-WT CFTR and corrected CFBE‑F508del CFTR cells, PLCb3 silencing also inhibits CFTR‑dependent current activated by forskolin and UTP-activated calcium-dependent chloride channels (CaCC). Our study supports the importance of PLC in maintaining CFTR‑dependent chloride secretion over time, getting maximal CFTR-dependent current and increasing CaCC activation in bronchial epithelial cells.

中文翻译:

磷脂酶C控制人支气管上皮细胞中氯依赖的短路电流

气道上皮细胞分泌的氯化物对于水和离子的动态平衡以及气道表面预防感染至关重要。这种分泌在几种人类疾病中都受到损害,包括囊性纤维化,一种由于CFTR基因突变导致氯离子通道缺陷的遗传病理。一种潜在的治疗方法旨在通过纠正突变的CFTR本身或通过刺激质膜上的非CFTR氯离子通道来增加氯离子的分泌。在这里,我们研究了磷脂酶C在调控人呼吸道上皮16HBE140和CFBE细胞中过表达WT-或F508del-CFTR的跨上皮氯化物分泌中的作用。蛋白质印迹分析显示三种内源性PLC亚型PLCd1,PLCg1和PLCb3在16HBE14o细胞中的表达。在16HBE14o细胞中,我们在沉默每种PLC同工型或使用PLC抑制剂U73122或其非活性类似物U73343之后进行了Ussing室实验。我们的结果表明,PLCb3和PLCg1与毛喉素激活的CFTR相关的短路电流有关,而与PLCd1无关。在CFBE-WT CFTR和校正后的CFBE-F508del CFTR细胞中,PLCb3沉默还抑制了由毛喉素和UTP激活的钙依赖性氯离子通道(CaCC)激活的CFTR依赖性电流。我们的研究支持PLC在维持CFTR依赖的氯化物随时间的分泌,获得最大CFTR依赖的电流以及增加支气管上皮细胞中CaCC活化方面的重要性。我们的结果表明,PLCb3和PLCg1与毛喉素激活的CFTR相关的短路电流有关,而与PLCd1无关。在CFBE-WT CFTR和校正后的CFBE-F508del CFTR细胞中,PLCb3沉默还抑制了由毛喉素和UTP激活的钙依赖性氯离子通道(CaCC)激活的CFTR依赖性电流。我们的研究支持PLC在维持CFTR依赖的氯化物随时间的分泌,获得最大CFTR依赖的电流以及增加支气管上皮细胞中CaCC活化方面的重要性。我们的结果表明,PLCb3和PLCg1与毛喉素激活的CFTR相关的短路电流有关,而与PLCd1无关。在CFBE-WT CFTR和校正后的CFBE-F508del CFTR细胞中,PLCb3沉默还抑制了由毛喉素和UTP激活的钙依赖性氯离子通道(CaCC)激活的CFTR依赖性电流。我们的研究支持PLC在维持CFTR依赖的氯化物随时间的分泌,获得最大CFTR依赖的电流以及增加支气管上皮细胞中CaCC活化方面的重要性。
更新日期:2020-11-25
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