Wang Y N, Pei C, Qin L, Li J M, Yi J L, Chen L
Department of Ophthalmology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, China.
Zhonghua Yan Ke Za Zhi. 2016 Apr 11;52(4):285-90. doi: 10.3760/cma.j.issn.0412-4081.2016.04.011.
To investigate the expression of transcription factors snail and slug in epithelial mesenchymal transition (EMT) of human lens epithelial cells (HLEC) induced by transforming growth factor-β2 (TGF-β2).
Experimental research. HLEC were treated with different concentrations of TGF-β2 (1.0 and 10.0 μg/L) for different time. The morphological changes were observed under inverted microscope. The expression and cellular localization of snail and slug were evaluated by immunofluorescence. Expressions of snail, slug, E-Cadherin and α-SMA were further determined by Western blot analysis. Single factor analysis of variance, rank sum test and Pearson correlation were used for statistical analysis.
Cultured HLEC were polygonal monolayer cells with tight intercellular adhesion closely and patchy distribution. After treatment of different doses of TGF-β2 for 24 h, HLEC became isolated, exhibited long spindle-like shape as fibroblastic phenotype. The immunofluorescence staining indicated that snail and slug were localized in the nuclei. The expressions of snail and slug appeared to be positive correlative to TGF-β2 dose (snail protein expression: 0.74±0.16, 1.13±0.03, 1.54±0.18 and slug protein expression: 1.96±0.02, 3.12±0.09, 4.07±0.12 in HLEC treated with 0.1, 1.0 and 10 μg/L TGF-β2 respectively) (χ(2)=9.62,P=0.022;F=241.10,P<0.01). In addition, the expression of α-SMA and E-Cadherin showed the similar form (α-SMA protein expression: 0.87±0.04, 1.42±0.11, 2.17±0.36 and E-Cadherin protein expression: 2.50±0.36, 1.65±0.32, 0.41±0.14 in HLEC treated with 0.1, 1.0 and 10.0 μg/L TGF-β2 respectively) (χ(2)=9.97,P=0.019;F=19.99,P<0.01). All Pearson correlation coefficient were close to 1. The expression of snail and slug in HLEC were also increased with extending duration of TGF-β2 (1.0 μg/L). The expression levels of both proteins were modestly up-regulated at 8 hours, robustly increased at 24 h, reached peak at 48h and began to decline at 72 h (snail protein expression: 0.90±0.13, 1.43±0.14, 1.96±0.27, 1.57±0.16 and slug protein expression: 0.91±0.36, 1.24±0.16, 2.44±0.26, 1.43±0.16 in HLEC treated with 1.0 μg/L TGF-β2 for 8 h, 24 h, 48 h and 72 h respectively) (F=12.49,P=0.001;F=14.03,P<0.01).
Transcription factors snail and slug might be time and dose-dependently involved in in-vitro TGF-β2-induced EMT of HLEC. (Chin J Ophthalmol, 2016, 52: 285-290).
探讨转录因子蜗牛（Snail）和蛞蝓（Slug）在转化生长因子-β2（TGF-β2）诱导的人晶状体上皮细胞（HLEC）上皮-间质转化（EMT）中的表达。
实验研究。用不同浓度（1.0和10.0μg/L）的TGF-β2处理HLEC不同时间。在倒置显微镜下观察形态变化。通过免疫荧光评估Snail和Slug的表达及细胞定位。进一步采用蛋白质印迹分析检测Snail、Slug、E-钙黏蛋白（E-Cadherin）和α-平滑肌肌动蛋白（α-SMA）的表达。采用单因素方差分析、秩和检验及Pearson相关性分析进行统计学分析。
培养的HLEC为多边形单层细胞，细胞间紧密黏附，分布成片。用不同剂量的TGF-β2处理24小时后，HLEC变得分散，呈现出成纤维细胞样的长梭形。免疫荧光染色显示Snail和Slug定位于细胞核。Snail和Slug的表达似乎与TGF-β2剂量呈正相关（分别用0.1、1.0和10μg/L TGF-β2处理的HLEC中，Snail蛋白表达：0.74±0.16、1.13±0.03、1.54±0.18；Slug蛋白表达：1.96±0.02、3.12±0.09、4.07±0.12）（χ(2)=9.62，P=0.022；F=241.10，P<0.01）。此外，α-SMA和E-Cadherin的表达呈现类似形式（分别用0.1、1.0和10.0μg/L TGF-β2处理的HLEC中，α-SMA蛋白表达：0.87±0.04、1.42±0.11、2.17±0.36；E-Cadherin蛋白表达：2.50±0.36、1.65±0.32、0.41±0.14）（χ(2)=9.97，P=0.019；F=19.99，P<0.01）。所有Pearson相关系数均接近1。随着TGF-β2（1.0μg/L）作用时间延长，HLEC中Snail和Slug的表达也增加。两种蛋白的表达水平在8小时时适度上调，24小时时显著增加，48小时时达到峰值，72小时时开始下降（分别用1.0μg/L TGF-β2处理8小时、24小时、48小时和72小时的HLEC中，Snail蛋白表达：0.90±0.13、1.43±0.14、1.96±0.27、1.57±0.16；Slug蛋白表达：0.91±0.36、1.24±0.16、2.44±0.26、1.43±0.16）（F=12.49，P=0.001；F=14.03，P<0.01）。
转录因子Snail和Slug可能在体外TGF-β2诱导的HLEC EMT中呈时间和剂量依赖性参与。（《中华眼科杂志》，2016年，52：285 - 290）
