ADAR1 shRNA对人胶质瘤U87细胞增殖和凋亡的影响

Effects of shRNA-ADAR1 on the Proliferation and Apoptosis of U87 Glioma Cell

目的:研究ADAR1 shRNA对人胶质瘤细胞U87细胞增殖和凋亡的影响。方法:通过构建ADAR1-shRNA的干扰质粒,经脂质体法转染胶质瘤U87细胞系,通过荧光倒置显微镜观察转染效率,选择转染效率最高的细胞系。取转染48h细胞,采用RT-PCR和Western-blot分别检测ADAR1 mRNA及蛋白的表达,流式细胞仪检测其细胞凋亡率,MTT法检测细胞增殖情况。结果:①经ADAR1-shRNA转染48h后的转染效率最高,此时U87细胞系中ADAR1 mRNA及蛋白的表达均被显著抑制,较阴性对照组及空白组均明显降低(P<0.05)。②在转染ADAR1-shRNA后,细胞凋亡率为(28.14%±3.76%),明显高于阴性对照组(3.20%±1.57%)和空白组(2.80%±1.49%),细胞增殖率较阴性对照组及空白组明显下降(P<0.05)。结论:通过shRNA抑制ADAR1的表达能明显促进人胶质瘤细胞U87细胞的凋亡和抑制其增殖,ADAR1基因可能成为治疗治疗胶质瘤的新靶点。

Objective： Thisstudywasdesigned to investigate the effects of transfection withsmall hairpinRNA（shRNA）-DAR1 onthe proliferation and apoptosis of human glioma U87 cell line. Methods： The shRNA expression vector which expresses the specific shRNA targeting ADAR1 mRNA （ADARI-hRNA） or independent sequence （Negative-hRNA） was transfected into U87 cells, and then the U87 cells with stable expression of shRNA were selected. The expressions of ADAR1 were detected by RT-CR and Western blotting. The cellular proliferation activity, and the apoptotic rate were determined by MTT assay and flow cytometry, respectively. Results： Compared with the Negative-hRNA group and the non-ransfection group, the mRNA and protein expression levels of ADAR1 in U87 cells with transfection of ADARI-hRNA （interference group） was significantly suppressed and the cell proliferation was slowed down after transfection for 48h （P〈0.05）. The apoptotic rate in the interference group （28.14%±3.76%） was significantly higher than those in the Negative-hRNA group （3.20%± 1.57%） and the non-ransfection group （2.80%± 1.49%） （P〈0.05）. The proliferation ratio of U87 cells significantly decreased after transfection of ADAR1 shRNA （P〈0.05）. Conclusion： ADARI-hRNA transfection increases the apoptotic rate, inhibits the proliferation of U87 glioma cells, which provides a potential target in gene therapy for glioma.