真核表达survivin基因对曲古菌素A诱导卵巢癌细胞凋亡作用的影响
Effect of Eukaryotic Expression of survivin on Apoptosis of Ovarian Cancer Cell Induced by Trichostatin A
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摘要: 目的 探讨survivin 基因对曲古菌素A( TSA) 诱导卵巢癌细胞凋亡作用的影响。方法 (1) 将本实验室已构建成功的survivin 正义全长真核表达质粒(pcDNA 3. 1-urvivin), 经脂质体包裹转染人卵巢癌细胞A2780, 以转染pcDNA 3. 1空载体的A2780 细胞为对照。(2) RT2PCR 和Western blot 方法分别检测survivin mRNA 和蛋白质的表达。(3) MTT 比色法和流式细胞仪(FACS) 分别检测TSA 对两组细胞存活率和凋亡率的影响。(4) Western blot 检测TSA 作用下A2780 细胞中survivin 蛋白的表达变化。结果 (1) RT-PCR和Western blot 检测提示转染pcDNA 3. 1-urvivin 组中survivin mRNA 和蛋白质表达明显高于空载体组。(2) MTT 比色法和FACS 检测提示转染pcDNA 3. 1-urvivin 组细胞存活率明显高于空载体组,细胞凋亡率明显低于空载体组,差异有统计学意义( P<0. 05) 。pcDNA 3. 1-urvivin转染后的2780 细胞对TSA 的敏感性明显降低。(3) Western blot 检测提示survivin 的表达水平随着TSA 作用时间的延长而下降。结论 曲古菌素A 诱导卵巢癌细胞的凋亡作用可能与survivin 基因表达有关。Abstract: Objective To investigate the effect of survivin expression on apoptosis of ovarian cancer cells induced by Trichostatin A( TSA) . Methods (1) A2780 cells were t ransfected with pcDNA 3. 1-urvivin (full2length survivin cDNA) which has been successfully const ructed in our laboratory. A2780 Cells t ransfected with pcDNA 3. 1 were cultured as cont rols. (2) The expression of survivin mRNA and protein was detected by RT-PCRandWestern blot, respectively. (3) Two cell groups were t reated with TSA and analyzed for suvival and apoptotic rate by MTT assay and flow cytomet ry ( FACS) . (4) Expression of survivin protein was detected under TSA t reatment by Western blot . Results (1) The level of survivin mRNA and protein in t ransfected cells t ransfected with pcDNA 3. 1-survivin was significantly higher than that in cont rol cells detected by RT-PCR and Western blot . (2) The viable rate of cells t ransfected with pcDNA 3. 1-survivin was obviously higher than that of cont rol cells and the apoptotic rate was significantly lower than cont rol detected by MTT assay and FACS, respectively ( P < 0. 05) . The sensitivity of A2780 cells t ransfected with pcDNA 3. 1-survivin to TSA was greatly reduced. (3) The expression of survivin in A2780 cells obviously decreased with TSA incubation time. Conclusion Apoptosis of ovarian cancer cells induced by TSA may be related to the expression of survivin gene.
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