TGF-β1诱导人胃腺癌SGC-7901细胞建立EMT模型

目的 拟利用TGF-β1诱导人胃癌SGC-7901细胞发生上皮间质转化（EMT）,建立EMT模型.方法 不同浓度TGF-β1作用SGC-7901细胞24h,观察其形态变化,并采用CCK8、迁移试验、细胞侵袭试验和蛋白印迹等技术,检测TGF-β1对SGC-7901细胞增殖、迁移、侵袭能力的影响.结果 经10ng/ml TGF-β1诱导24h后,多数细胞变为梭型细胞边界基本消失,细胞间间隙明显增大.采用CCK-8试验检测不同浓度（0、5、10、20ng/ml）的TGF-β1对SGC-7901细胞作用24h后OD值,结果显示各作用浓度对SGC-7901的生长抑制率未发现有增加或减少趋势.体外划痕试验检测细胞迁移能力实验中,各组间距为94.67±3.06、87.00 ±3.61、68.67±7.77和91.00±3.00（单位：像素）,10ng/ml剂量组有大量细胞迁移至划痕区,划痕间距明显变小（P＜0.05）.Transwell小室侵袭实验结果：0、5、10、20ng/ml组,侵袭的细胞数分别为152.00±5.29、169.00±6.00、299.33±10.50和176.00±11.27,且l0ng/ml剂量组细胞数明显增多（P＜0.05）.蛋白免疫印迹定量分析显示：不同浓度（0、5、10、20ng/ml）的TGF-β1作用SGC-7901细胞24h,E-cadherin蛋白灰度值测量结果：0、5、10、20ng/ml组,分别为0.95 ±0.02、0.91±0.04、0.62±0.05和0.75±0.32,10ng/ml剂量组灰度值明显减少（P＜0.05）;Vimentin蛋白灰度值测量结果：0、5、10、20ng/ml组,分别为1.18±0.03、1.33±0.02、1.57±0.05和1.19±0.07,10ng/ml剂量组灰度值明显增加（P＜0.05）.结论 10ng/ml TGF-β1诱导人胃腺癌SGC-7901细胞24h后,细胞明显发生EMT过程,本研究证明在人胃腺癌SGC-7901细胞系,通过10ng/ml TGF-β1的诱导24h可以成功建立EMT模型.

By TGF-b1 Induced Human Gastric Cancer SGC-7901 Cells Establish EMT Model

Objective By using TGF - β1 induced human gastric cancer SGC - 7901 cells which occurs epithelial - mesenchymal transformation and establish the model of the epithelial - mesenchymal transition （ EMT）. Methods Using the different concentrations of TGF - β1 effect on SGC - 7901 ceils for 24h, observing the morphological changes, using CCK8, migration assay, cell invasion assay and Western blot techniques to test TGF - β1 on SGC -7901 cell＇s capacity of proliferation, migration, and invasion. Results After induc- tion by lOng/mL TGF - β1 in 24h, the majority of cells turned to fusiform cell, cell borders disappeared, the gap between cells had sig- nificantly increasing. After impact of different concentrations of TGF - β1 on SGC - 7901 cells in 24h, SGC - 7901 cells OD values checked by CCK -8 assay showed that the inhibition of SGC -7901 growth rate didn＇t increase or decrease. In the experiments of chec- king cell migration capacity by the vitro scratch test, the group spacing were ： 94.67± 3.06, 87.00 ± 3.61, 68.67 ± 7.77 and 91. 00 ± 3.00 （ units pixels）, a large number of cell of 10ng/ml group migrated to the scratch area, scratch spacing was significantly smaller （ P 〈 0. 05）. Transwell chamber invasion assay results： 0, 5, 10, 20ng/ml group, the corresponding number of cell invasion were 152.00± 5.29, 169.00±6.00, 299.33 ± 10.50 and 176.00± 11.27, and the 10ng/ml group＇s number of cells increased significantly （P 〈 0.05 ）. Quantitative of Western blot analysis showed that ： after different concentrations （0,5,10,20ng/ml） of TGF - β1 induce SGC - 7901 cells 24h, E - cadherin protein gray value measurements ： 0,5,10,20ng/ml groups were 0.95 ±0.02,0.91 ± 0.04,0.62 ± 0.05 and 0. 75 ± 0.32, 10ng/ml group＇s gray values significantly decreased （P 〈 0.05）; Vimentin protein gray value measurements showd that 0,5,10, 20ng/ml group were 1.18 ± 0.03,1.33 ±0. 02,1.57 ± 0. 05 and 1.19± 0.07, 10ng/ml group＇s gray values increased sig-nificantly （P〈0.05）. Conclusion Cells SGC -7901 inducted by 10ng/ml TGF - β1 in 24h, occurs EMT significantly. This study demonstrates that： using human gastric carcinoma cell line SGC -7901 induced by 10ng/ml TGF -β1 in 24h, EMT model can be successfully established.