Snail基因沉默对胰腺癌PANC1细胞侵袭和增殖的影响

The effect of Snail gene silencing on cell invasion and proliferation in human pancreatic cancer cell line PANC1

目的观察Snail基因沉默后对胰腺癌PANC1细胞侵袭和增殖能力的影响。方法构建针对Snail的小发卡RNA（shRNA-Snail）的慢病毒载体和不针对任何已知mRNA序列的shRNA（shRNA-NC）慢病毒载体，分别感染PANC1细胞，以未感染细胞作为对照组。应用实时荧光定量PCR法和蛋白质印迹法检测Snail、α-平滑肌肌动蛋白（α-SMA）及E-钙黏素（E-cadherin）mRNA和蛋白的表达；Transwell小室检测细胞体外侵袭能力，CCK-8法检测细胞增殖能力。结果与shRNA-NC组比较，shRNA-Snail组细胞Snail mRNA和蛋白表达水平显著下降[（0.27±0.02）比（0.92±0.03），（0.26±0.02）比（0.80±0.02）]，α-SMA mRNA和蛋白表达水平亦显著下降[（0.33±0.04）比（0.97±0.07），（0.31±0.04）比（0.74±0.06）]，E-cadherin mRNA和蛋白表达水平则显著升高[（1.57±0.45）比（0.95±0.08），（0.86±0.03）比（0.20±0.03）]，穿膜细胞数显著减少[（6.80±0.73）比（26.80±2.52）个/400倍视野]，细胞增殖明显被抑制[（0.74±0.05）比（1.47±0.04）]，差异均有统计学意义（P值均〈0.01）。shRNA-NC组与对照组细胞各指标的差异均无统计学意义。结论沉默Snail基因表达可在一定程度上抑制胰腺癌PANC1细胞的侵袭和增殖能力。

Objective To observe the effect of silencing Snail gene on the invasion and proliferation ability of human pancreatic cancer cell line PANC1. Methods Lentiviral vectors that can express small hairpin RNA（shRNA） targeting human Snail gene（shRNA-Snail） or shRNA sequence that did not match any known mRNA（shRNA-NC） were constructed, and transfected into PANC1 cells. Untransfected cells served as control, mRNA and protein expression of Snail, or-smooth muscle actin （α-SMA） and E-cadherin was determined by real time quantitative PCR and Western blotting, respectively. In vitro invasion ability was tested by Transwell model. Proliferation ability was measured by CCK-8 assay. Results Compared with those in shRNA-NC group, Snail mRNA （ 0.27 ± 0.02 vs 0.92 ± 0.03 ） and protein level （ 0. 26 ± 0. 02 vs 0. 80 ± 0.02） ,and α-SMA mRNA （0.33 ±0.04 vs 0.97 ±0.07） and protein level （0.31 ±0.04 vs 0.74 ±0.06） in shRNA-Snail group were obviously decreased, but E-cadherin mRNA （1.57 ± 0.45 vs 0. 95 ±0. 08 ） and protein level （0.86 ± 0.03 vs 0. 20 ± 0.03 ） were greatly increased. The number of cells permeating the septum of transwell [ （ 6.80 ± 0.73 ）/400 magnification vs （ 26.80 ± 2.52 ）/400 magnification, P 〈0.01 ] was significantly decreased, and cell proliferation was inhibited（ 0.74 ± 0. 05 vs 1.47 ± 0.04, P 〈 0.01 ）. All the differences above were statistically significant （ all P 〈 0.01 ）. No significant differences were observed between shRNA-NC and normal control group. Conclusions Silencing Snail gene may restrain the invasion and proliferation ability of PANC1 ceils to a certain degree.