TGF-β1对HLEC发生EMT影响分析

The effect of transforming growth factor-β1 on epithelial-mesenchymal transition in human lens epithelial cells

目的研究TGF-β1对人晶状体上皮细胞(HLEC)上皮间质转分化(EMT)的影响。方法根据处理不同,将细胞分为TGF-β1组、siRNA-TGF-β1组和空白对照组,培养24 h后观察细胞形态,Transwell法检测各组细胞迁移能力,实时荧光定量PCR检测各组细胞中TGF-β1、α-SMA、E-cadherin和vimentin基因表达,Western blotting法检测各组细胞中PI3K、Akt和p-Akt蛋白表达。结果 TGF-β1组迁移细胞数为(31.2±4.7)个,高于siRNA-TGF-β1组(14.6±5.3)个及空白对照组(3.9±1.4)个,且空白对照组高于siRNA-TGF-β1组;siRNA-TGF-β1组TGF-β1、α-SMA及Vimentin的mRNA相对表达量均低于TGF-β1组及空白对照组,且空白对照组低于TGF-β1组;siRNA-TGF-β1组E-cadherin的mRNA相对表达量均高于TGF-β1组及空白对照组,且空白对照组高于TGF-β1组;siRNA-TGF-β1组细胞中PI3K蛋白及p-Akt蛋白相对表达量均低于TGF-β1组和空白对照组,且空白对照组低于TGF-β1组,siRNA-TGF-β1组Akt蛋白相对表达量高于TGF-β1处理组和空白对照组,且空白对照组高于TGF-β1组。结论 TGF-β1可诱导HLEC发生EM T,且可促进HLEC迁移,其机制可能与细胞中PI3K/Akt信号通路活化有关。

Objective To investigate the effect of transforming growth factor （TGF）-β1 on the epithelial-mesenchymal transition （EMT） in human lens epithelial cells （HLECs）. Methods HLECs were cultured separately, according their treatment group. For this purpose, cells were divided into the TGF-β1 treatment group, siRNA-TGF-β1 group, and the control group. After culturing cells for 24 h in their respective treatment groups, cell morphologies were observed. Cell migration in each treatment group was detected using the transwell migration assay. The expression levels of TGF-β1, α-SMA, E-cadherin, and vimentin genes were detected using real-time polymerase chain reaction. The expression lev- els of PI3K, Akt, and p-Akt proteins were detected using Western blotting. Results In the TGF-β1 treatment group, 31.2 ± 4.7 HLECs were found to have transformed. This number was significantly higher than that in the siRNA-TGF- β1 （ 14.6 ± 5.3 ） and control groups （3.9 ± 1.4）. Specifically, the number of cells transformed in the siRNA-TGF-β1 group was higher than that in the control group. Expression levels of TGF-β1 mRNA, α-SMA mRNA, and vimentin mRNA were the highest in the TGF-β1 treatment group, followed by lower levels in the siRNA-TGF-β1 group, and the lowest levels in the control group. In contrast, the expression levels of E-cadherin mRNA were highest in the siRNA- TGF-β1 group, followed by lower levels in the control group and the lowest levels in the TGF-β1 treatment group. These differences were statistically significant （P 〈 0.05 ）. The expression levels of PI3K and p-Akt proteins were the highest in the cells of the TGF-β1 treatment group, followed by lower levels in the control group. These levels were lowest in the siRNA-TGF-β1 group. The relative expression levels of Akt proteins were the highest in the siRNA-TGF- β1 group, followed by lower levels in the control group and the lowest levels in the TGF-β1 treatment group; these differences were statistically significant （ P 〈 0.05 ）. Conclusion TGF-β1 could induce HLECs towards EMT, and thus promote HLEC migration. This mechanism could be related to the activation of PI3K/Akt signaling pathway.