Congmin Gu, Mu Zeng, Yihui Ma, Hanliang Lin
Department of Pathology, Guangzhou Women and Children Hospital, Guangzhou, PR China.
Leuk Lymphoma. 2006 Sep;47(9):1941-8. doi: 10.1080/10428190600725354.
RNA interference (RNAi) has been widely used in tumor gene therapy, antivirus and gene drug selection. Survivin gene is highly expressed in non-Hodgkin's lymphoma (NHL) tissues and high malignancy Burkitt's lymphoma cell line-Daudi and it is regarded as a potential target of gene therapy for NHL. This study used a vector-based short hairpin RNA (shRNA) technique to explore the effect of RNAi-mediated survivin gene silencing on apoptosis and proliferation of Daudi cells. Recombinant plasmid survivin-shRNA was transfected into Daudi cells transiently and stably. The expression of survivin was detected by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The apoptosis of Daudi cells after transfection were evaluated by flow cytometry. After transfection of survivin-shRNA, the levels of survivin mRNA were significantly reduced by 64.20% (transient transfection) and 62.32% (stable transfection), respectively; The levels of survivin protein were significantly reduced by 63.50% (transient transfection) and 61.88% (stable transfection); compared with control-shRNA and PBS treated groups. Apoptosis of Daudi cells were significantly higher in the transfection group than in the control group, respectively 21.30 +/- 2.96% (transient transfection) and 19.10 +/- 2.15% (stable transfection). In conclusion, it was suggested that survivin could be an attractive target for new anti-cancer intervention of NHL and vector-based survivin-shRNA could effectively reduce the expression of survivin and induce cell apoptosis and growth inhibition of NHL cells.
RNA干扰（RNAi）已广泛应用于肿瘤基因治疗、抗病毒及基因药物筛选。生存素基因在非霍奇金淋巴瘤（NHL）组织及高恶性伯基特淋巴瘤细胞系——Daudi中高表达，被视为NHL基因治疗的潜在靶点。本研究采用基于载体的短发夹RNA（shRNA）技术，探讨RNAi介导的生存素基因沉默对Daudi细胞凋亡及增殖的影响。将重组质粒生存素-shRNA瞬时及稳定转染至Daudi细胞。通过半定量逆转录-聚合酶链反应（RT-PCR）及蛋白质免疫印迹法检测生存素的表达。采用流式细胞术评估转染后Daudi细胞的凋亡情况。转染生存素-shRNA后，生存素mRNA水平分别显著降低64.20%（瞬时转染）和62.32%（稳定转染）；生存素蛋白水平分别显著降低63.50%（瞬时转染）和61.88%（稳定转染），与对照-shRNA及PBS处理组相比。转染组Daudi细胞的凋亡率显著高于对照组，分别为21.30±2.96%（瞬时转染）和19.10±2.15%（稳定转染）。综上所述，提示生存素可能是NHL新型抗癌干预的有吸引力的靶点，基于载体的生存素-shRNA可有效降低生存素表达，诱导NHL细胞凋亡并抑制其生长。
