siRNA干扰MACC1基因对食管癌细胞TE-1的生物学影响

Biological effects of MACC1 siRNA interference on esophageal carcinoma TE-1 cells

目的通过siRNA干扰沉默结肠癌转移相关基因1（metastasis．associatedincoloncanceY1，MACCI），观察其对食管癌细胞株TE-1增殖、迁移及侵袭能力的影响。方法通过化学合成的特异性siRNA与阳离子脂质体Lipofectamine TM 2000形成复合体，转染食管癌细胞株TE-1，应用qRT-PCR、Westernblot检测转染后MACCl基因和蛋白表达水平；MTT法检测细胞生长增殖水平；划痕实验、Transwell实验检测细胞株干扰前后迁移侵袭能力的改变。结果siRNA．MACCl转染TE一1细胞后，与空白组比较，干扰组MACCl基因和蛋白的表达水平明显降低，分别降低71％和87％；MTT实验观察1—7d，干扰组较空白组、阴性对照组增殖速度明显降低（P〈0．05），第4天后效果更明显；划痕实验结果显示干扰组迁移划痕处的细胞（10．6±2．2）明显少于空白组（68．2±2．5），差异具有统计学意义（P〈0．05），各对照组间无统计学差异（P〉0．05）；Transwell实验结果显示干扰组穿过膜的TE-1细胞数目（4．2±1．9）较空白组（58．4±7．4）明显减少（P〈0．05），各对照组间无统计学差异（P〉0．05），表明干扰后TE-1细胞的迁移侵袭能力显著降低。结论siRNA．MACCl干扰导致TE-1细胞MACCl基因及蛋白表达水平明显下调；MACCl基因下调后，食管癌TE-1细胞的增殖、迁移、侵袭能力均明显降低，说明MACCl基因具有促进食管癌细胞增殖、迁移、侵袭的能力。

Objective To determine the effects of siRNA interfering metastasis-associated in colon cancer 1 （MACC1） on the extracorporeal proliferation, migration and invasion of the esophageal squamous cell carcinoma （ESCC） cell line TE-1. Methods The special MACCI-siRNA sequence was chemically synthesize, and then transfected into the TE-1 cells with lipofectamineTM 2000. RT-quantity PCR and Western blotting were used to detect the mRNA and protein level of MACC1 in untreated TE-1 cells, lipo2000-treated cells （mock）, siRNA-nc transfected cells, and siRNA-MACC1 transfected cells. MTI＂ assay was used to detect the proliferation of TE-1 ceils. Scratching test and Transwell chamber assay were evaluated the migration of the cells. Results Compared with the untreated cell, mock, and siRNA-nc cells, the expression of MACC1 was obviously decreased by 71% at mRNA and 87% at protein levels. In 7 dafter transfection, MTT assay indicated that the proliferation of the transfected TE-1 ceils was significantly inhibited compared with the other 3 types of ceils （ P 〈 0.05 ）, especially from the fourth day. Scratching test showed that there were obviously less migrated cells from the siRNA-MACC1 group than those from the control （ 10.6 ± 2.2 vs 68.2 ± 2.5, P 〈 0.05 ）. Transwell assay also showed the invasion ability of TE-1 ceils transfected with siRNA-Maccl was lesser than the control cells （4.2 ± 1.92 vs 58.4 ± 7.44, P 〈 O. 05 ）. But no significant difference was seen among the untreated cell, mock, and siRNA-nc cells. Conclusion siRNA interference of MACC1 obviously down-regulates the molecule at mRNA and protein levels in TE-1 cells, and results in significant inhibition in the proliferation, migration and invasion of the cells. Therefore, MACC1 might be involved in the growth, migration and invasion of ESCC.