目的：应用RNA干扰技术下调肺癌A549细胞中Slug基因的表达，探讨其对A549细胞增殖、细胞周期和细胞侵袭能力的影响。方法：构建靶向Slug基因的shRNA真核表达质粒pGPU6-GFP-Neo-Slug，与阴性对照质粒pNeg-shRNA分别用脂质体法转染A549细胞。Real-time PCR和Western blotting验证转染后Slug mRNA和蛋白的表达。CCK-8法、流式细胞术和Transwell实验分别检测下调Slug表达对A549细胞增殖、细胞周期和侵袭能力的影响。结果：成功构建pGPU6-GFP-Neo-Slug载体并转染A549细胞，转染率达90%。与pNeg-shRNA组和空白对照组相比，pSlug-shRNA组A549细胞中Slug mRNA\[（0.23±0.01）vs（0.97±0.08）、（1.0±0.09），P＜0.05\]和蛋白\[（0.20±0.09 ）vs（1.0±0.32）、（1.13±0.26），P＜0.05\]表达量显著降低；细胞增殖抑制率明显上升\[（35.3±5.4）% vs（1.5±0.2）%、（3.3±0.7）%，均P< 0.01\]；细胞增殖指数显著降低 \[（32.92±0.69）% vs（48.19±0.71）%、（42.88±0.75）%，均P<0.05\]，并且处于G1期细胞数明显增多\[（67.08±092）% vs（52.81±0.78）%、（56.12±0.73）%，均P<0.05\]；细胞侵袭能力显著降低\[穿透基底膜细胞数：（55±9）vs（169±12）、（173±15），均P<0.01\]。结论: pGPU6-GFP-Neo-Slug转染肺癌A549细胞能有效下调Slug基因的表达，从而抑制A549的增殖侵袭能力、阻滞细胞周期于G1期。

Objective:To study the effect of silencing Slug gene on proliferation, cell cycle and invasion ability of lung cancer A549 cells by RNA interference technique. Methods: The recombinant plasmid expressing short hairpin RNA (shRNA) targeting Slug gene was constructed and named as pGPU6-GFP-Neo-Slug. A549 cells were transfected with pGPU6-GFP-Neo-Slug and the negative control plasmid pNeg-shRNA with liposome method. Real-time PCR and Western blotting analysis was performed to determine the expression of Slug mRNA and protein after transfection, respectively. The proliferation, cell cycle distribution and cell invasion of A549 cells were detected by CCK-8 assay, flow cytometry and transwell assay, respectively. Results: pGPU6-GFP-Neo-Slug vector was successfully constructed and transfected into A549 cells with transfection rate of nearly 90%. Compared to the control and pNeg-shRNA group, Slug mRNA (\[0.23±0.01\] vs \[0.97±0.08\], \[1.0±0.09\]; all P＜0.05) and protein (\[0.20±0.09\] vs \[1.0±0.32\], \[1.13±0.26\]; all P＜0.05) expression level was significantly reduced in pSlug-shRNA group. Compared to the control and pNeg-shRNA group, the inhibition rate of proliferation in Slug-silencing A549 cells was significantly increased (\[35.3±5.4\]% vs \[15±02%\], \[3.3±0.7%\]; all P< 0.05); the cell multiplication index was significantly decreased(\[32.92±069\] vs \[48.19±0.71\], \[42.88±0.75\]; all P<0.05); the cell number in G1 phase was significantly increased(\[67.08±092\] vs \[52.81±0.78\], \[56.12±0.73\]; all P<0.05); and the cell invasion ability of A549 was significantly reduced (number of alive A549 cells through the matrigel chamber：\[55±9\] vs \[169±12\], \[173±15\]; all P<0.01). Conclusion: pGPU6-GFP-Neo-Slug vector transfected into lung cancer A549 cell can effectively silence Slug gene expression, inhibit cell proliferation, influence cell cycle and inhibit cell invasion of A549 cells.

国家自然科学基金资助项目（No.81102057）