High glucose-induced epithelial-mesenchymal transition contributes to the upregulation of fibrogenic factors in retinal pigment epithelial cells
High glucose-induced epithelial-mesenchymal transition contributes to the upregulation of fibrogenic factors in retinal pigment epithelial cells
复制标题
高葡萄糖诱导的上皮间质转化有助于视网膜色素上皮细胞中纤维形成因子的上调。
DOI：
10.3892/ijmm.2016.2768
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发表时间：
2016-12-01
影响因子：
5.4
通讯作者：
Gao, Guoquan
中科院分区：
文献类型：
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作者：
Che, Di;Zhou, Ti;Gao, Guoquan
据报道，上皮间质转化（EMT）介导多种生理和病理过程。然而，高糖诱导的视网膜色素上皮细胞（RPE细胞）EMT的发生及其致病作用尚不清楚。本研究的目的是检查高葡萄糖对 RPE 细胞 EMT 的影响。将培养的 RPE 细胞暴露于 25 mM D-葡萄糖。将编码Snail基因的载体和靶向Snail的siRNA（Snail siRNA）转染至细胞中，分别诱导Snail的过表达或沉默。 AKT 和细胞外信号调节激酶 (ERK) 抑制剂分别用于阻断 AKT 和 ERK 的激活。通过蛋白质印迹分析和免疫荧光染色测定EMT标志物、纤维形成因子、磷酸化ERK和磷酸化AKT的水平。通过伤口愈合测定评估细胞迁移。我们的结果表明，高葡萄糖提高了关键的 EMT 转录因子 Snail 和其他间充质标记物的表达，并促进细胞迁移。此外，Snail的过表达会升高细胞中纤连蛋白和结缔组织生长因子（CTGF）的水平，而Snail的沉默会降低细胞中高糖诱导的纤连蛋白和CTGF的表达。从机制上讲，AKT抑制剂（AKT抑制剂IV）和ERK抑制剂（U0126）显着降低了Snail的表达，以及高糖诱导的纤连蛋白和CTGF的水平。总体而言，据我们所知，本研究首次证明了高糖诱导的RPE细胞间充质标志物的上调，并提示间充质转化可能参与视网膜疾病的病理过程。
It has been reported that epithelial-mesenchymal transition (EMT) mediates Multiple physiological and pathological processes. However, the occurrence and the pathogenic role of high glucose-induced EMT in retinal pigment epithelial cells (RPE cells) is unknown. The aim of this study was to examine the effects of high glucose on EMT in RPE cells. Cultured RPE cells were exposed to 25 mM D-glucose. A vector encoding the Snail gene and siRNA targeting Snail (Snail siRNA) were transfected into the cells to induce the overexpression or silencing of Snail, respectively. AKT and extracellular signal-regulated kinase (ERK) inhibitors were used to block the activation of AKT and ERK, respectively. The levels of EMT markers, fibrogenic factors, phosphorylated ERK and phosphorylated AKT were determined by western blot analysis and immunofluorescence staining. Cell migration was evaluated by wound healing assay. Our results revealed that high glucose elevated the expression of the key EMT transcriptional factor, Snail, and that of other mesenchymal makers, and promoted cell migration. Moreover, the overexpression of Snail elevated the levels of fibronectin and connective tissue growth factor (CTGF), whereas the silencing of Snail decreased the expression of fibronectin and CTGF induced by high glucose in the cells. Mechanistically, the AKT inhibitor (AKT inhibitor IV) and ERK inhibitor (U0126) significantly decreased the expression of Snail, as well as the levels of fibronectin and CTGF which were induced by high glucose. On the whole, and to the best of our knowedge, the present study is the first to demonstrate the upregulation of mesenchymal markers in RPE cells induced by high glucose, and suggest that mesenchymal transition may be involved in the pathological processes of retinal diseases.