Upregulation of Connexin43 Expression in Corneal Fibroblasts by Corneal Epithelial Cells
Upregulation of Connexin43 Expression in Corneal Fibroblasts by Corneal Epithelial Cells
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DOI：
10.1167/iovs.08-2418
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发表时间：
2009-05-01
影响因子：
4.4
通讯作者：
Nishida, Teruo
中科院分区：
文献类型：
--
作者：
Ko, Ji-Ae;Yanai, Ryoji;Nishida, Teruo
目的.作者先前已经表明，角膜上皮细胞中紧密连接蛋白的表达受共培养系统中角膜成纤维细胞的调节。他们现在已经研究了角膜上皮细胞对角膜成纤维细胞中缝隙连接蛋白connexin 43表达的影响。将人角膜成纤维细胞和猿猴病毒40转化的人角膜上皮（HCE）细胞培养在胶原玻璃胶膜的相对侧上。通过逆转录-聚合酶链反应、免疫印迹和免疫荧光分析检测角膜成纤维细胞中连接蛋白的表达。胰岛素样生长因子-1（IGF-1）mRNA特异性小干扰RNA对连接蛋白43表达的影响通过转染确定。角膜成纤维细胞中连接蛋白43 mRNA和蛋白的量增加HCE细胞的存在。HCE细胞对角膜成纤维细胞中紧密连接蛋白ZO-1、occludin和claudin的表达没有影响。通过将角膜成纤维细胞暴露于IGF-1来模拟HCE细胞对连接蛋白43表达的影响。此外，通过RNA干扰去除HCE细胞中的IGF-1在很大程度上消除了这些细胞对角膜成纤维细胞中连接蛋白43表达的影响。最后，HCE细胞对角膜成纤维细胞中连接蛋白43表达的上调被丝裂原活化蛋白激酶ERK（PD 98059）和p38 MAPK（SB 203580）信号传导抑制剂阻断。角膜上皮细胞的存在上调了角膜成纤维细胞中连接蛋白43的表达，表明角膜上皮细胞对于维持角膜成纤维细胞之间的缝隙连接介导的通讯是重要的。（Invest Ophthalmol维斯科学。2009;50：2054-2060）DOI：10.1167/iovs.08-2418
PURPOSE. The authors have previously shown that the expression of tight-junction proteins in corneal epithelial cells is regulated by corneal fibroblasts in a coculture system. They have now examined the effect of corneal epithelial cells on the expression of the gap-junction protein connexin43 in corneal fibroblasts.METHODS. Human corneal fibroblasts and simian virus 40 transformed human corneal epithelial (HCE) cells were cultured on opposite sides of a collagen vitrigel membrane. Expression of junctional proteins in corneal fibroblasts was examined by reverse transcription-polymerase chain reaction, immunoblot, and immunofluorescence analyses. The effect of a small interfering RNA specific for insulin-like growth factor-1 (IGF-1) mRNA on connexin43 expression was determined by transfection.RESULTS. The amounts of connexin43 mRNA and protein in corneal fibroblasts were increased by the presence of HCE cells. HCE cells had no effect on the expression of the tight-junction proteins ZO-1, occludin, and claudin in corneal fibroblasts. The effect of HCE cells on connexin43 expression was mimicked by exposure of corneal fibroblasts to IGF-1. Furthermore, depletion of IGF-1 in HCE cells by RNA interference largely abolished the effect of these cells on connexin43 expression in corneal fibroblasts. Finally, the upregulation of connexin43 expression in corneal fibroblasts by HCE cells was blocked by inhibitors of signaling by the mitogen-activated protein kinases ERK (PD98059) and p38 MAPK (SB203580).CONCLUSIONS. The presence of corneal epithelial cells upregulated the expression of connexin43 in corneal fibroblasts, suggesting that corneal epithelial cells are important for the maintenance of gap junction-mediated communication among corneal fibroblasts. (Invest Ophthalmol Vis Sci. 2009;50:2054-2060) DOI:10.1167/iovs.08-2418