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目的研究RNA干扰技术(RNAi)能否失活Rad51基因,从而使肿瘤细胞的放射敏感性增加。方法应用Psilencer U6-1．0质粒为载体在人纤维肉瘤HT1080细胞内表达短链干扰RNA (siRNA)。RNAi效应在蛋白水平用Western blot方法评价；在细胞水平用成克隆分析及放射诱导的凋亡来评价。结果所选3个靶序列均起到失活Rad51的作用,放射诱导的Rad51高表达也能被RNAi抑制。成克隆分析表明实验组存活分数在照射2、4、6、8Gy后,分别是对照组的80．5%、78．6%、69．0%、57．7%(P=0．020)；实验组放射诱导的凋亡在放射后0、6、18、24、48h分别是对照组的1．03、1．43、1．19、1．29、1．33倍(P=0．017)。结论RNAi技术能够使内源性Rad51基因及放射诱导的Rad51高表达失活,从而使人纤维肉瘤HT1080细胞放射敏感性增加；RNAi技术是一种新的研究基因功能和放射增敏的方法。
Objective To investigate whether RNA interference (RNAi) can inactivate Rad51 gene and thus increase the radiosensitivity of tumor cells. Methods Psilencer U6-1.0 plasmid was used as a vector to express short-chain interfering RNA (siRNA) in human fibrosarcoma HT1080 cells. RNAi effects were assessed by Western blot at the protein level; clonogenic assays and radiation-induced apoptosis at the cellular level were evaluated. Results The three selected target sequences all played the role of inactivating Rad51. Radradiation-induced high expression of Rad51 was also inhibited by RNAi. Clone analysis showed that the survival scores of experimental group were 80.5%, 78.6%, 69.0%, 57.7% (P = 0.020) after irradiation for 2, 4, 6 and 8 Gy respectively The apoptosis induced by radiation in the experimental group was 1.03, 1.43, 1.19, 1.29, 1.33 times higher than those in the control group at 0, 6, 18, 24 and 48 hours after radiation (P = 017). Conclusion RNAi can inactivate the endogenous Rad51 gene and radiation-induced high expression of Rad51, thereby increasing the radiosensitivity of human fibrosarcoma HT1080 cells. RNAi is a new method to study gene function and radiosensitization.