siRNA沉默survivin基因表达对恶性胶质瘤细胞放射敏感性的影响

目的探讨针对人survivin基因的siRNA（small interfering RNA）对恶性胶质瘤细胞放疗敏感性的影响。方法设计并合成针对survivin基因的siRNA片断,构建干涉质粒载体pSil4.1-shsurvivin1和pSil4.1-shsurvivin2（pSil4.1-shcontrol作为阴性对照）。利用脂质体辅助转染恶性胶质瘤细胞（U251）,以G418筛选稳定表达干涉载体的细胞系,RT-PCR和W estern blot试验检测survivin的m RNA和蛋白表达,并筛选survivin基因显著抑制的稳定转染细胞系（U251-s2）。通过克隆形成试验和皮下移植瘤试验分别在体外和体内检测恶性胶质瘤细胞放射敏感性的变化。结果相对于未转染和阴性对照组细胞,U251-s2细胞中survivin mRNA和蛋白表达分别显著下调了40.5%和51.8%;同时,survivin基因表达下调能够显著增强恶性胶质瘤细胞的体内外放射敏感性。结论Survivin基因过表达和恶性胶质瘤的放疗抗性密切相关,抑制其表达可以显著增强恶性胶质瘤细胞的放疗敏感性,具有潜在的临床应用前景。

Effects of siRNA-Mediated Survivin on Radiosensitivity of Glioblastoma

Objective To explore the effects of small interfering RNA（siRNA）targeting survivin on the radiosensitivity of glioblastoma cells.Methods siRNA vectors targeting survivin（pSil 4.1-shsurvivin 1 and pSil 4.1-shsurvivin 2）were successfully designed and constructed,while pSil 4.1-shcontrol was considered as negative control.Those vectors were transfected into glioblastoma cell（U251）and the stably transfected cells were also selected by G418.RT-PCR and Western blot assays were performed to detect the expression of survivin mRNA and protein.The inhibition effects of survivin on in vitro and in vivo radiosensitivity of U251 cells were detected by colony formation assay and animal experiment.Results Compared with untransfected and negative control U251 cells,the levels of survivin mRNA and protein expression in U251-s2 cells stably expressing shsurvivin2 were significantly reduced by 40.5%and 51.8%,respectively.Moreover,inhibition of survivin could significantly enhance radiosensitivity of U251 cells both in vitro and in vivo.Conclusion The overexpression of survivin gene is correlated with radioresistance of glioblastoma cells.Thus,inhibition of survivin expression could significantly enhance the radiosensitivity of glioblastoma cell and might be a potential strategy for the treatment of glioblastoma.