Connexin 26 expression prevents down-regulation of barrier and fence functions of tight junctions by Na+/K+-ATPase inhibitor ouabain in human airway epithelial cell line Calu-3
Connexin 26 expression prevents down-regulation of barrier and fence functions of tight junctions by Na+/K+-ATPase inhibitor ouabain in human airway epithelial cell line Calu-3
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DOI：
10.1016/j.yexcr.2006.08.014
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发表时间：
2006-11-15
影响因子：
3.7
通讯作者：
Sawada, Norimasa
中科院分区：
文献类型：
--
作者：
Go, Mitsuru;Kojima, Takashi;Sawada, Norimasa
间隙连接被认为在上皮细胞的分化中发挥着至关重要的作用，并且与紧密连接蛋白相关。在本研究中，为了研究间隙连接在调节紧密连接的屏障功能和栅栏功能中的作用，我们将Cx26基因引入人气道上皮细胞系Clau-3中，并使用Na+/K+-ATP酶抑制剂哇巴因构建紧密连接破坏模型。在亲本 Calu-3 细胞中，RT-PCR 检测到间隙连接蛋白 Cx32 和 Cx43，但未检测到 Cx26，以及紧密连接蛋白 occludin、JAM-1、ZO-1、claudin-1、-2、-3、-4、-5、-6、-7、-8、-9 和 -14。紧密连接的屏障功能和栅栏功能保持良好，而GJIC水平较低。哇巴因治疗导致紧密连接的屏障功能和栅栏功能破坏，同时 occludin、JAM-1、claudin-2 和 -4 下调，ZO-1 和 claudin-14 上调。在Cx26转染子中，通过Western blotting和免疫细胞化学检测到Cx26蛋白，并且观察到许多间隙连接斑块和发育良好的紧密连接链。与亲本细胞相比，Cx26 转染子中claudin-14 的表达显着增加，并且在一些细胞中，Cx26 与claudin-14 共定位。有趣的是，用 Cx26 转染可防止哇巴因处理对紧密连接两种功能的破坏，而不会改变紧密连接蛋白。用 GJIC 阻滞剂 18 甘草次酸和油酰胺预处理不影响 Cx26 转染引起的变化。这些结果表明，Cx26 表达（而不是介导的细胞间通讯）可能调节人气道上皮细胞系 Calu-3 中的紧密连接屏障和栅栏功能。 (c) 2006 Elsevier Inc. 保留所有权利。
Gap junctions are considered to play a crucial role in differentiation of epithelial cells and to be associated with tight junction proteins. In this study, to investigate the role of gap junctions in regulation of the barrier function and fence function on the tight junctions, we introduced the Cx26 gene into human airway epithelial cell line Clau-3 and used a disruption model of tight junctions employing the Na+/K+-ATPase inhibitor ouabain. In parental Calu-3 cells, gap junction proteins Cx32 and Cx43, but not Cx26, and tight junction proteins occludin, JAM-1, ZO-1, claudin-1, -2, -3, -4, -5, -6, -7, -8, -9, and -14 were detected by RT-PCR. The barrier function and fence function of tight junctions were well maintained, whereas the GJIC was low level. Treatment with ouabain caused disruption of the barrier function and fence function of tight junctions together with down-regulation of occludin, JAM-1, claudin-2, and -4 and up-regulation of ZO-1 and claudin-14. In Cx26 transfectants, Cx26 protein was detected by Western blotting and immunocytochemistry, and many gap junction plaques were observed with well-developed tight junction strands. Expression of claudin-14 was significantly increased in Cx26 transfectants compared to parental cells, and in some cells, Cx26 was co-localized with claudin-14. Interestingly, transfection with Cx26 prevented disruption of both functions of tight junctions by treatment with ouabain without changes in the tight junction proteins. Pretreatment with the GJIC blockers 18 glycyrrhetinic acid and oleamide did not affect the changes induced by Cx26 transfection. These results suggest that Cx26 expression, but not the mediated intercellular communication, may regulate tight junction barrier and fence functions in human air-way epithelial cell line Calu-3. (c) 2006 Elsevier Inc. All rights reserved.