Alcohol increases the permeability of airway epithelial tight junctions in Beas-2B and NHBE cells.
Alcohol increases the permeability of airway epithelial tight junctions in Beas-2B and NHBE cells.
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DOI：
10.1111/j.1530-0277.2011.01640.x
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发表时间：
2012-03
期刊：
影响因子：
--
通讯作者：
Sisson JH
中科院分区：
文献类型：
--
作者：
Simet SM;Wyatt TA;DeVasure J;Yanov D;Allen-Gipson D;Sisson JH
紧密连接在细胞之间形成连续的带状结构，并用于调节细胞旁信号传导。蛋白激酶C（PKC）已被证明可以调节紧密连接的组装和拆卸，并被酒精激活。先前的研究表明，酒精会增加肺泡细胞紧密连接的渗透性。然而，很少有人知道酒精的影响，在上皮细胞的紧密连接进行气道。我们假设，长期酒精暴露减少zonnula occluden-1（ZO-1）和claudin-1定位在细胞膜上，并通过PKC依赖性机制增加渗透性。为了验证这一假设，我们将正常人支气管上皮细胞（NHBE）、来自人支气管上皮转化细胞系（Beas-2B）的细胞和表达PKCα显性阴性（DN）的Beas-2B暴露于酒精（20、50和100 mM）长达48小时。免疫荧光用于评估ZO-1、密蛋白-1、密蛋白-5和密蛋白-7定位的变化。电细胞基质阻抗传感（ECIS）用于测量NHBE、Beas-2B和DN细胞的单层之间的紧密连接的渗透性。乙醇以浓度依赖性方式增加紧密连接的通透性，并降低ZO-1，claudin-1，claudin-5和claudin-7在细胞膜上的定位。为了确定可能的信号传导机制，我们测量了PKC亚型（α，δ，β，ζ）的活性。100 mM酒精作用1-6 h后，Beas-2B细胞中PKCα活性显著升高，而PKC β活性在1 h后显著降低，3 h后显著升高。用Gö-6976抑制PKCα可以防止酒精诱导的细胞膜上ZO-1和claudin-1的蛋白质变化。PKCα显性阴性的Beas-2B细胞对酒精诱导的蛋白质改变具有抵抗性。这些结果表明，酒精通过激活PKCα破坏ZO-1、claudin-1、claudin-5和claudin-7，导致酒精诱导的支气管上皮细胞“渗漏”。这种酒精诱导的气道渗漏状态可能导致与急性和慢性酒精摄入相关的气道宿主防御受损。
Tight junctions form a continuous belt-like structure between cells and act to regulate paracellular signaling. Protein kinase C (PKC) has been shown to regulate tight junction assembly and disassembly and is activated by alcohol. Previous research has shown that alcohol increases the permeability of tight junctions in lung alveolar cells. However, little is known about alcohol’s effect on tight junctions in epithelium of the conducting airways. We hypothesized that long-term alcohol exposure reduces zonnula occluden-1 (ZO-1) and claudin-1 localization at the cell membrane and increases permeability through a PKC-dependent mechanism. To test this hypothesis, we exposed normal human bronchial epithelial cells (NHBE), cells from a human bronchial epithelial transformed cell line (Beas-2B), and Beas-2B expressing a PKCα dominant negative (DN) to alcohol (20, 50, and 100 mM) for up to 48 hours. Immunofluorescence was used to assess changes in ZO-1, claudin-1, claudin-5 and claudin-7 localization. Electrical cell substrate impedance sensing (ECIS) was used to measure permeability of tight junctions between monolayers of NHBE, Beas-2B, and DN cells. Alcohol increased tight junction permeability in a concentration-dependent manner and decreased ZO-1, claudin-1, claudin-5 and claudin-7 localization at the cell membrane. To determine a possible signaling mechanism, we measured the activity of PKC isoforms (alpha, delta, epsilon, zeta). PKCα activity significantly increased in Beas-2B cells from 1–6 hours of 100 mM alcohol exposure, while PKCζ activity significantly decreased at 1 hour and increased at 3 hrs. Inhibiting PKCα with Gö-6976 prevented the alcohol-induced protein changes of both ZO-1 and claudin-1 at the cell membrane. PKCα dominant negative Beas-2B cells were resistant to alcohol-induced protein alterations. These results suggest that alcohol disrupts ZO-1, claudin-1, claudin-5 and claudin-7 through the activation of PKCα, leading to an alcohol-induced “leakiness” in bronchial epithelial cells. Such alcohol-induced airway leak state likely contributes to the impaired airway host defenses associated with acute and chronic alcohol ingestion.
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DOI：
10.1152/ajpcell.1997.273.3.c937
发表时间：
1997-09-01
影响因子：
5.5
作者：
Liedtke, CM;Cole, T;Ikebe, M
通讯作者：
Ikebe, M
影响因子：
2.3
作者：
Fernandez, Alberto L.;Koval, Michael;Guidot, David M.
通讯作者：
Guidot, David M.
影响因子：
3.7
作者：
Clarke, H;Ginanni, N;Mullin, JM
通讯作者：
Mullin, JM
影响因子：
4.9
作者：
Asgrimsson, V;Gudjonsson, T;Baldursson, O
通讯作者：
Baldursson, O
影响因子：
3.3
作者：
Fahy, J V;Corry, D B;Boushey, H A
通讯作者：
Boushey, H A