Guan Hai-Tao, Xue Xing-Huan, Wang Xi-Jing, Li Ang, Qin Zhao-Yin
Department of Oncological Surgery, Second Affiliated Hospital, Medical College of Xi'an Jiaotong University, Xi'an 710004.
Chin Med Sci J. 2006 Jun;21(2):115-9.
To construct an expression vector of small interfering RNA (siRNA) against survivin and observe its effects on survivin expression and proliferation of human pancreatic cancer cell line PC-2 and breast cancer cell line MCF-7.
Constructed an expression vector of siRNA against survivin and transfected it into PC-2 and MCF-7 cells using lipofectamine 2000. The expression of survivin was detected by semi-quantitive RT-PCR and immunohistochemistry, and its effects on proliferation of PC-2 and MCF-7 cells were detected by MTT assay.
The introduction of sequence-specific siRNA could efficiently suppress survivin expression at both mRNA and protein levels in the two cancer cell lines. In PC-2 cells, the expression inhibition rates were 81.25% at mRNA level and 74.24% at protein level. In MCF-7 cells, the expression inhibition rates were 64.91% at mRNA level and 79.72% at protein level. The proliferation of PC-2 and MCF-7 cells was also suppressed, and 24 and 48 hours after the cells were reseeded, the proliferation inhibition rates of PC-2 cells were 28.00% and 33.38%, and that of MCF-7 cells were 31.58% and 33.02%, respectively.
The expression vector of siRNA against survivin can block survivin expression in PC-2 and MCF-7 cells efficiently and specifically. Down regulation of survivin expression can suppress proliferation of PC-2 and MCF-7 cells. Survivin RNAi may have potential value in gene therapy of human cancers.
构建针对生存素(survivin)的小干扰RNA(siRNA)表达载体,并观察其对人胰腺癌细胞系PC-2和乳腺癌细胞系MCF-7中survivin表达及细胞增殖的影响。
构建针对survivin的siRNA表达载体,并用脂质体2000将其转染至PC-2和MCF-7细胞。采用半定量逆转录聚合酶链反应(RT-PCR)和免疫组织化学法检测survivin的表达,用噻唑蓝(MTT)比色法检测其对PC-2和MCF-7细胞增殖的影响。
导入序列特异性siRNA可有效抑制两种癌细胞系中survivin在mRNA和蛋白水平的表达。在PC-2细胞中,mRNA水平的表达抑制率为81.25%,蛋白水平为74.24%。在MCF-7细胞中,mRNA水平的表达抑制率为64.91%,蛋白水平为79.72%。PC-2和MCF-7细胞的增殖也受到抑制,细胞重新接种后24小时和48小时,PC-2细胞的增殖抑制率分别为28.00%和33.38%,MCF-7细胞的增殖抑制率分别为31.58%和33.02%。
针对survivin的siRNA表达载体可有效、特异地阻断PC-2和MCF-7细胞中survivin的表达。survivin表达下调可抑制PC-2和MCF-7细胞的增殖。Survivin RNA干扰在人类癌症的基因治疗中可能具有潜在价值。