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目的采用RNA干扰技术探讨生存素(survivin)对人肺腺癌细胞增殖及凋亡的影响及其可能的分子机制。方法采用脂质体介导将pGCsi-sh survivin与pGCsi质粒转染人肺腺癌细胞株A549,经G418筛选获得稳定抑制survivin表达的肺腺癌细胞模型;Western blot检测转染细胞survivin、p21、鼠双微体基因2(MDM2)蛋白表达;MTT比色法检测阳性细胞克隆的增殖活性;流式细胞仪检测各组细胞凋亡。结果成功构建稳定抑制survivin表达的肺腺癌细胞模型。与对照组相比,反义组p21表达及细胞凋亡率显著增加,而MDM2表达及细胞增殖显著降低。结论下调A549细胞survivin表达后,细胞增殖降低而细胞凋亡增加,其作用机制与p21表达增加和MDM2降低有关。
Objective To investigate the effect of survivin on the proliferation and apoptosis of human lung adenocarcinoma cells and its possible molecular mechanism by RNAi technique. Methods The human lung adenocarcinoma cell line A549 was transfected with pGCsi-sh survivin and pGCsi by lipofectamine. The lung adenocarcinoma cells stably expressing survivin were screened by G418. The expression of survivin, p21, The expression of MDM2 protein was detected by MTT assay. The proliferation of positive clone was detected by MTT colorimetric assay. Apoptosis was detected by flow cytometry. Results The lung adenocarcinoma cell model which stably inhibited the expression of survivin was constructed successfully. Compared with the control group, p21 expression and apoptosis rate of antisense group increased significantly, while MDM2 expression and cell proliferation decreased significantly. Conclusion The down-regulation of survivin expression in A549 cells decreases the proliferation of cells and increases the apoptosis of A549 cells. Its mechanism is related to the increase of p21 expression and the decrease of MDM2.