Wang H F, Yang H, Hu L B, Lei Y H, Qin Y, Li J, Bi C W, Wang J S, Huo Q
Department of Urology, The Second Affiliated Hospital of Kunming Medical University, Kunming, Yunnan, China.
Department of Urology, Yunnan Provincial Tumor Hospital, The Third Affiliated Hospital of Kunming Medical University, Kunming, Yunnan, China
Genet Mol Res. 2014 Nov 27;13(4):9939-50. doi: 10.4238/2014.November.27.23.
We investigated the effect of siRNA targeting enhancer of EZH2 on cell proliferation, invasion, migration, and apoptosis of human bladder cancer T24 cells. An siRNA-expressing plasmid targeting the EZH2 gene was transfected into T24 cells. Quantitative polymerase chain reaction and Western blot analysis were used to detect EZH2 expression at the mRNA and protein levels, respectively. Proliferation, invasion, and migration of T24 cells were examined in vivo using MTT, wound healing, and transwell chamber migration assays, respectively. Annexin V-fluorescein isothiocyanate/propidium iodide flow cytometric analysis was performed to determine cell apoptosis levels. Expression of EZH2 in T24 cells was suppressed at the mRNA and protein levels. Following transfection for 48 h, growth was inhibited by 37.9%, which was markedly lower than that in the negative control group (P < 0.05). Following a wound-healing assay for 24 h, transfected cell migration distance was 1.37 ± 0.12, which was markedly less than the horizontal migration distance of negative control group cells (P < 0.01). In addition, the cell invasion ability of EZH2- siRNA group cells decreased by 67% compared with negative control group cells (P < 0.01). Following transfection for 48 h, early- and late-stage apoptosis rates for T24 cells were 22.8 and 3.60%, respectively, which were higher than in the negative control group (P < 0.01). EZH2 gene silencing effectively suppressed the proliferation, invasion, and migration abilities of human bladder cancer cells, promoting apoptosis.
我们研究了靶向EZH2增强子的小干扰RNA(siRNA)对人膀胱癌T24细胞增殖、侵袭、迁移和凋亡的影响。将靶向EZH2基因的表达siRNA的质粒转染到T24细胞中。分别采用定量聚合酶链反应和蛋白质印迹分析检测EZH2在mRNA和蛋白质水平的表达。分别使用MTT、伤口愈合和Transwell小室迁移试验在体内检测T24细胞的增殖、侵袭和迁移。进行膜联蛋白V-异硫氰酸荧光素/碘化丙啶流式细胞术分析以确定细胞凋亡水平。T24细胞中EZH2的表达在mRNA和蛋白质水平均受到抑制。转染48小时后,生长受到37.9%的抑制,明显低于阴性对照组(P<0.05)。伤口愈合试验24小时后,转染细胞的迁移距离为1.37±0.12,明显小于阴性对照组细胞的水平迁移距离(P<0.01)。此外,与阴性对照组细胞相比,EZH2-siRNA组细胞的细胞侵袭能力下降了67%(P<0.01)。转染48小时后,T24细胞的早期和晚期凋亡率分别为22.8%和3.60%,高于阴性对照组(P<0.01)。EZH2基因沉默有效抑制了人膀胱癌细胞的增殖、侵袭和迁移能力,促进了细胞凋亡。