siRNA 下调Gli 基因影响食管腺癌OE33 细胞生长转移的实验研究

Silencing Gli gene with siRNA modulates growth and invasion of esophageal adenocarcinoma OE33 cells

摘要:

目的 研究Gli 表达下调对食管腺癌OE33 细胞生长转移的影响，并探讨其相关的分子机制。 方法 将Gli1、Gli2 siRNA 和空白对照siRNA 转染OE33 细胞48 h，实时荧光定量聚合酶链反应检测OE33 细胞Gli1 和Gli2 mRNA 表达水平；采用Western blot 检测OE33 细胞中Gli1、Gli2、Cyclin D1、p27、 E-cadherin 及N-cadherin 蛋白的表达水平；流式细胞术检测OE33 细胞增殖和细胞周期分布；Transwell 实 验检测细胞侵袭能力。结果 采用Gli1 和Gli2 siRNA 抑制Gli1 和Gli2 mRNA 及蛋白表达后，可下调Cyclin D1 和N-cadherin 蛋白表达，增加E-cadherin 和p27 蛋白表达，同时抑制细胞增殖，将细胞周期阻滞在G0/G1 期并降低细胞的侵袭能力，与对照组比较，差异有统计学意义（P <0.05）。结论 Gli 在食管腺癌的生长转移 中具有重要作用，敲掉Gli 1 和Gli 2 基因表达可抑制食管腺癌细胞周期分布和上皮- 间质转化能力，这可能与 Cyclin D1、p27、E-cadherin 及N-cadherin 蛋白变化相关。

Abstract:

Objective To explore the effect of down-regulation of Gli expression on growth and invasion of esophageal adenocarcinoma OE33 cells and to discuss the molecular mechanisms. Methods Gli1 and Gli2 siRNA and control siRNA were respectively transfected into OE33 cells for 48 h. Expressions of the Gli1 and Gli2 mRNA and protein were detected by real-time fluorescence quantitative PCR and Western blot respectively. The expressions of cyclin D1, p27, E-cadherin and N-cadherin proteins were observed by Western blot. The cell cycle was determined by flow cytometry. In addition, invasion ability was detected by Transwell assay. Results After transfection of Gli1 and Gli2 siRNA, Gli1 and Gli2 expressions were inhibited, the expressions of cyclin D1 and N-cadherin proteins were downregulated, and E-cadherin and p27 expressions were increased (P < 0.05). After Gli1 and Gli2 were knocked down, cell cycle was arrested at G0/G1 phase and cell invasion ability was decreased compared with the control siRNA (P < 0.05). Conclusions Gli may play an important role in the growth and invasion of esophageal adenocarcinoma. Gli1 and Gli2 knockout can inhibit the cell cycle and epithelial-mesenchymal transition, which may be associated with cyclin D1, p27, E-cadherin and N-cadherin.