摘要
目的 探讨转录因子Snail及Slug在转化生长因子-β2（TGF-β2）诱导的体外培养人晶状体上皮细胞（HLEC）上皮间质转分化（EMT）中的表达.方法 实验研究.采用不同浓度的TGF-β2作用于HLEC一定时间后,倒置荧光显微镜观察细胞形态变化;免疫荧光法观察转录因子Snail及Slug细胞内的表达及分布情况;Western blot测定Snail,Slug,E-cadherin及α-SMA蛋白的表达水平.实验结果采用单因素方差分析、秩和检验及Pearson相关性检验.结果 体外培养的HLEC呈多角形单层细胞,细胞间粘连紧密,呈片状分布,加入不同浓度TGF-β2继续培养24 h后,细胞转变为长梭形类成纤维细胞样形态,并移行为单个细胞分布;免疫荧光提示Snail及Slug蛋白的表达位于细胞核;TGF-β2诱导Snail及Slug的表达呈现时间和浓度依赖性,不同浓度（0.1、1.0、10.0 μg/L）的TGF-β2处理HLEC24 h后,各实验组Snail （0.74 ±0.16、1.13±0.03、1.54±0.18）、Slug（1.96±0.02、3.12±0.09、4.07±0.12）、α-SMA （0.87±0.04、1.42±0.11、2.17±0.36）和E-Cadherin（2.50±0.36、1.65±0.32、0.41±0.14）蛋白及对照组的表达差异有统计学意义（x2 =9.62,P=0.02;F=241.10,P＜0.01;x2=9.97,P=0.02;F=19.99,P＜0.01）,并且Snail、Slug分别与α-SMA和E-Cadherin的表达变化呈现高度相关性（相关系数均接近1,P＜0.01）.用1μg/L的TGF-β2处理HLEC不同时间（8、24、48、72 h）后发现8h即有Snail和Slug蛋白的表达上调,24 h表达量上调明显,48 h表达最高,72 h表达量开始下降.各实验组Snail蛋白（0.90±0.13、1.43±0.14、1.96±0.27、1.57±0.16）和Slug蛋白（0.91±0.36、1.24±0.16、2.44±0.26、1.43±0.16）的表达及对照组差异有统计学意义（F=12.49,P=0.001;F=14.03,P＜0.01）.结论 转录因子Snail及Slug可能参与TGF-β2诱导的体外HLEC的EMT过程,且呈浓度与时间依赖性.
Objective 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）.Methods 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.Results 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） （x2=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） （x2=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 （1.0μg/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 �
出处
《中华眼科杂志》
CAS
CSCD
北大核心
2016年第4期285-290,共6页
Chinese Journal of Ophthalmology
基金
国家自然科学基金（81470614,81300786）
陕西省国际交流项目（2012-KW-31）
江西省自然科学基金（20132BAB205024）
高等学校博士学科点专项科研基金新教师类（20133601120012）
关键词
晶体
上皮细胞
上皮.间质转化
转录因子
转化生长因子Β2
Lens, crystalline
Epithelial cells
Epithelial-mesenchymal transition
Transcription factors
Transforming growth factor beta2
