RNA干扰介导的胶质瘤细胞IGF-1R基因下调诱导肿瘤细胞凋亡  

Down-regulation of IGF-1R gene expression by RNA interference induces glioma cells apoptosis

作　　者：胡锦[1] 王元元[2] 董万利[2] 蒋觉安[2] 金由辛[3] 

机构地区：[1]上海交通大学附属第六医院神经外科,上海200233 [2]苏州大学附属第一医院神经内科 [3]上海中国科学院生化所分子生物学国家重点实验室

出　　处：《中国神经精神疾病杂志》2006年第3期224-227,共4页Chinese Journal of Nervous and Mental Diseases

基　　金：国家自然科学基金(编号:30371459);上海市科学技术发展基金(编号:034047)资助

摘　　要：目的应用RNA干扰(RNAi)技术介导胶质瘤C6细胞株胰岛素生长因子1受体(IGF1R)基因下调诱导C6细胞凋亡的研究。方法体外化学合成靶向IGF1R基因的小干扰RNA(siRNA),脂质体法将siRNA以不同浓度梯度转染C6细胞,设非特异的siRNA阳性对照组和未转染siRNA的阴性对照组:同时使用绿色荧光素标记的小干扰RNA(FITCsiRNA)转染细胞,荧光显微镜下观察siRNA转染效率;RT-PCR法半定量检测siRNA对IGF1R基因表达的抑制作用;流式细胞术检测siRNA诱导细胞凋亡作用。结果荧光显微镜下荧光素标记的siRNA转染组可见细胞质内清晰的绿色荧光,脂质体OligofectamineTM2000的转染效率接近100%;化学合成的siRNA明显抑制IGF1RmRNA的表达,各特异性siRNA不同浓度转染组IGF1RmRNA表达水平可下调约10%～80%,流式细胞术检测细胞凋亡率转染组为22.47%±3.15%,与阳性对照组2.3%±0.9%和阴性对照组1.14%±0.79%比较有明显提高,而阳性、阴性对照组相比无明显差异。结论化学合成的靶向IGF1R的siRNA可以明显下调IGF1R基因的表达,胶质瘤细胞凋亡率明显上升,为进一步进行胶质瘤的基因治疗研究奠定基础。Objective To study the inhibitory effect of RNA interference on glioma IGF-1R gene expression in addition to the apoptotic induction. Methods Small interference RNAs （si RNAs）targeting IGF-1R gene were synthesized chemically in vitro. C6 cells stably expressing IGF-1R gene were transfected with the siRNA by using oligofetamine^TM 2000 reagent , both the nontransfected cells and nonspecific siRNAs transfected cells were taken as negative and positive controls, siRNA transfection efficiency was detected by fluorescent microscopy, down-regulation of IGF-1R was demonstrated by RTPCR,apoptosis of C6 cell was measured by flow cytometry. Results Green fluorescence in the cells was seen clearly in FITC-siRNA transfected group under the fluorescent microscope, the transfection efficiency of oligofectamine^TM 2000 was approximately 100%. The expression of IGF-1R mRNA decreased by 10% - 80% in the siRNAs transfected C6 cells by RTPCR scan analysis as compared with that of the controls. Cell apoptosis could be induced from 1.14% ± 0. 79% in control group to 22. 47% ± 3. 15% in IGF1R-siRNA transfected group. Conclusions At the cell level, the chemically synthesized siRNAs targeted IGF-1R gene could down -regulate the expression of IGF-1R. In addition, a remarkable apoptosis was induced when compared with the control cell line . It was suggested that the suppression of IGF-1R expression and the induction of apoptotic provide the new method for the study of glioma gene therapy.

关 键 词：小干扰RNA 胰岛素样生长因子-1受体 胶质瘤 细胞凋亡 

分 类 号：R739.4[医药卫生—肿瘤]