微小RNA-1297对胃癌细胞侵袭迁移和Notch/zeb1信号通路的影响

Effect of microRNA-1297 on invasion/migration and Notch/zeb1 signaling pathway of gastric cancer cells

目的探讨微小RNA-1297(miR-1297)对胃癌细胞侵袭迁移和Notch/zeb1信号通路的影响。方法采用实时荧光定量PCR(QPCR)检测正常胃黏膜细胞GES-1和胃癌细胞株AGS、MKN45、HGC27及SGC7901中miR-1297的表达水平;选取miR-1297表达量最低的胃癌细胞分别转染miR-1297模拟物(miR-1297组)和阴性对照(miR-NC组),QPCR检测各组的miR-1297表达水平,划痕实验和Transwell小室实验分别检测迁移和侵袭能力,Western blotting检测上皮间质转化过程及Notch/zeb1信号通路中相关蛋白的表达。结果 GES-1细胞中miR-1297表达水平为1.01±0.05,高于AGS、MKN45、HGC27和SGC7901细胞的0.44±0.03、0.54±0.05、0.61±0.06和0.51±0.08,差异有统计学意义(P<0.05),选取AGS细胞进行后续实验。miR-1297组的miR-1297表达水平较miR-NC组升高(2.77±0.33)倍(P=0.018)。转染48 h后miR-1297组的划痕愈合率为(32.1±4.3)%,低于miR-NC组的(68.7±5.1)%,miR-1297组的侵袭细胞数目为(27±2)个,低于miR-NC组的(52±4)个,以上差异均有统计学意义(P<0.05)。Western blotting结果显示,与miR-NC组比较,miR-1297组E-钙黏蛋白水平升高(P<0.05),而N-钙黏蛋白、波形蛋白、Notch和zeb1水平均降低(P<0.05)。结论胃癌细胞中miR-1297水平降低。上调miR-1297表达可抑制胃癌细胞的迁移侵袭能力,可能与抑制Notch/zeb1信号通路有关。

Objective To investigate the effect of microRNA-1297 (miR-1297) on invasion/migration and Notch/zeb1 signaling pathway of gastric cancer cells. Methods Real-time quantitative PCR (QPCR) was used to detect the expression of miR-1297 in normal gastric mucosal cells GES-1 and gastric cancer cell lines AGS, MKN45, HGC27 and SGC7901. Gastric cancer cells with the lowest expression of miR-1297 were transfected with miR-1297 mimics (miR-1297 group) and negative control (miR-NC group), respectively. The expression of miR-1297 in each group was detected by QPCR. Scratch assay and Transwell chamber assay were used to detect the migrative and invasive ability of both groups, respectively. Western blotting was used to detect the expression of related proteins in the process of epithelial mesenchymal transformation and Notch/zeb1 signaling pathway. Results The expression level of miR-1297 in GES-1 cells was 1.01±0.05, significantly higher than that in AGS, MKN45, HGC27 and SGC7901 cells (0.44±0.03, 0.54±0.05, 0.61±0.06 and 0.51±0.08). The difference was statistically significant( P < 0.05). AGS cells were selected for subsequent experiments. The expression level of miR-1297 in miR-1297 group was (2.77±0.33) folds higher than that in miR-NC group ( P = 0.018). After 48 h, the healing rate of scratches in the miR-1297 group was (32.1±4.3)%, significantly lower than (68.7±5.1)% in miR-NC group ( P < 0.05). The number of invasive cells in the miR-1297 group was 27±2, significantly lower than 52±4 in the miR-NC group ( P < 0.05). Compared with the miR-NC group, the expression of E-cadherin increased in the miR-1297 group ( P < 0.05), while the expression of N-cadherin, Vimentin, Notch and zeb1 decreased ( P < 0.05). Conclusion The level of miR-1297 in gastric cancer cells decreased. Upregulation of the expression of miR-1297 can inhibit the migration and invasion of gastric cancer cells, which may be related to the inhibition of Notch/zeb1 signaling pathway.