Pannexin 1 Contributes to ATP Release in Airway Epithelia
Pannexin 1 Contributes to ATP Release in Airway Epithelia
复制标题
DOI：
10.1165/rcmb.2008-0367oc
复制
发表时间：
2009-11-01
影响因子：
6.4
通讯作者：
Salathe, Matthias
中科院分区：
文献类型：
--
作者：
Ransford, George A.;Fregien, Nevis;Salathe, Matthias
ATP是气道上皮细胞关键功能的旁分泌调节剂，但其释放机制尚不清楚。Pannexin （Panx）蛋白，与无脊椎动物的innexins相关，形成通道（称为pannexon），能够从几种细胞类型中释放ATIP。因此，在气道上皮细胞中通过泛子检测ATP释放。定量RT-PCR显示，正常人气道上皮细胞在气液界面（ALI）再分化过程中表达Panx1，表达水平与肺泡巨噬细胞相当；Panx3未表达。免疫组化显示Panx1在气道上皮的顶极表达。暴露于低渗应激下的ALI培养物在激发后1分钟内释放ATP的估计最大值为255 (+/- 64)nM （n = 6个来自三个不同肺部的培养物）或约1.5 (+/- 0.4)μ M（重新计算为正常气道表面液体体积）。使用日期和培养匹配的细胞（每个n >= 16来自4个不同的肺），pannexon抑制剂carbenoxolone （10 μ M）和probenecid (11 μ M)，而不是连接子抑制剂氟芬那酸（100 μ M），抑制ATP释放约60%。这些药物相应地影响了外源性Panx1在爪蟾卵母细胞中的电流。此外，利用慢病毒介导的shRNA在分化的气道上皮细胞中抑制Panx1的表达，也能抑制低渗应激时ATP的释放约60%。这些数据不仅表明Panx1在分化的气道上皮细胞中顶端表达，而且还有助于这些细胞中ATP的释放。
ATP is a paracrine regulator of critical airway epithelial cell functions, but the mechanism of its release is poorly understood. Pannexin (Panx) proteins, related to invertebrate innexins, form channels (called pannexons) that are able to release ATIP from several cell types. Thus, ATP release via pannexons was examined in airway epithelial cells. Quantitative RT-PCR showed Panx1 expression in normal human airway epithelial cells during redifferentiation at the air-liquid interface (ALI), at a level comparable to that of alveolar macrophages; Panx3 was not expressed. Immunohistochemistry showed Panx1 expression at the apical pole of airway epithelia. ALI cultures exposed to hypotonic stress released ATP to an estimated maximum of 255 (+/- 64) nM within 1 minute after challenge (n = 6 cultures from three different lungs) or to approximately 1.5 (+/- 0.4) mu M, recalculated to a normal airway surface liquid volume. Using date- and culture-matched cells (each n >= 16 from 4 different lungs), the pannexon inhibitors carbenoxolone (10 mu M) and probenecid (11 mM), but not the connexon inhibitor flufenamic acid (100 mu M), inhibited ATP release by approximately 60%. The drugs affected Panx1 currents in Xenopus oocytes expressing exogenous Panx1 correspondingly. In addition, suppression of Panx1 expression using lentivirus-mediated production of shRNA in differentiated airway epithelial cells inhibited ATP release upon hypotonic stress by approximately 60% as well. These data not only show that Panx1 is expressed apically in differentiated airway epithelial cells but also that it contributes to ATP release in these cells.