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目的:探讨RNA干扰(RNAi)技术沉默Bmi-1基因表达对人胃癌细胞株AGS增殖和侵袭能力的影响。方法:利用慢病毒表达体系pHelper1.0/pHelper2.0/pGCL-GFP,构建针对Bmi-1基因shRNA序列的RNAi重组质粒vshRNA-Bmi-1,转染AGS细胞,采用Western blot检测重组质粒对Bmi-1蛋白表达的影响,MTT法检测重组质粒对AGS细胞体外生长的抑制作用,PI单染法流式细胞术检测转染重组质粒后细胞凋亡与细胞周期的变化,小室侵袭实验检测AGS细胞侵袭能力变化。结果:成功构建了vshRNA-Bmi-1重组质粒,并成功转染AGS细胞抑制Bmi-1蛋白的表达。转染重组质粒后,AGS细胞增殖明显受到抑制,细胞凋亡增加,S期比例上升,细胞侵袭能力弱于非特异性转染组。结论:vshRNA-Bmi-1重组质粒明显下调Bmi-1蛋白在AGS胃癌细胞中的表达,并抑制肿瘤细胞的增殖与侵袭,促进其凋亡。
Objective: To investigate the effect of silencing Bmi-1 gene expression on the proliferation and invasion of human gastric cancer cell line AGS by RNA interference (RNAi) technique. Methods: RNAi recombinant plasmid vshRNA-Bmi-1 was constructed by lentivirus expression system pHelper1.0 / pHelper2.0 / pGCL-GFP and transfected into AGS cells. Western blot was used to detect the expression of Bmi The protein expression of AGS was detected by MTT assay. The inhibition of AGS cells in vitro was detected by MTT assay. The apoptosis and cell cycle of AGS cells were detected by flow cytometry. The cell invasion assay was used to detect AGS cells Invasive ability changes. Results: The vshRNA-Bmi-1 recombinant plasmid was successfully constructed and successfully transfected into AGS cells to inhibit the expression of Bmi-1 protein. After transfected with recombinant plasmid, AGS cell proliferation was significantly inhibited, apoptosis increased, the proportion of S phase increased, cell invasion was weaker than non-specific transfection group. Conclusion: The recombinant plasmid vshRNA-Bmi-1 significantly down-regulates the expression of Bmi-1 protein in AGS gastric cancer cells and inhibits the proliferation and invasion of tumor cells and promotes apoptosis.