siRNA对乳腺癌细胞CyclinE表达和生长抑制作用

Inhibition of Cyclin E Expression and Cell Proliferation by Small Interfering RNA in Breast Cancer MCF-7 Cell Line

研究siRNA对乳腺癌MCF-7细胞株cyclinE表达的抑制及对细胞生长的影响。化学合成针对cyclinE基因的小干扰RNA(siRNA),转染MCF-7细胞株;分别应用荧光定量PCR和免疫印迹测定cyclinEmRNA和蛋白质的表达,CCK-8测定细胞的增殖活性,流式细胞仪检测细胞周期,软琼脂培养检测细胞克隆形成能力。10、50、100nmol/LsiRNA-cyclinE分别使MCF-7细胞cyclinE基因表达降低了24.7%、62.5%和71.0%,蛋白质表达降低了40.8%、66.5%和71.3%。转染siRNA-cyclinE后,G1期细胞增多,S期减少,增殖受到抑制,软琼脂克隆形成率降低。结果提示,在MCF-7细胞株中,导入针对cyclinE的siRNA,可有效抑制cyclinE的表达,进而使细胞增殖减缓,逆转其恶性表型。

To study the inhibition of cyclin E expression by small interfering RNA （siRNA）, and alters cell proliferation in breast cancer MCF-7 cell line. The siRNA targeting cyclin E was chemically synthesized and transfected into MCF-7 cells by Oligofectamine, the expression of cyclin E was analyzed by quantitive PCR and Western blot, cell growth inhibition was measure with CCK-8 assay, cell cycle of the transfected cells was examined by flow cytometry, and cell colony forming ability was measured by soft-agar colony formation assay. After MCF- 7 cells were transfected with 10, 50, 100 nmol/L siRNA, the expression of cyclin E mRNA was respectively suppressed with inhibition rate of 24.7%, 62.5% and 71.0%, the corresponding decrease of cyclin E protein expression was confirmed; cells in G1 phase increased after transfecting with siRNA-cyclin E, but cells in S phase decreased; lacking cyclin E expression exhibited significantly inhibited cell proliferation and showed less colony forming ability in the breast cancer cell line MCF-7 Cells. These results indicate that siRNA-cyclin E was a useful tool for silencing cyclin E and inhibiting cell proliferation in breast cancer cell line.