沉默Survivin基因对鼻咽癌CNE1细胞增殖和凋亡的研究

Research on the Effect of Silence of Survivin Gene by RNAi on the Proliferation and Apoptosis of Nasopharyngeal Carcinoma CNE1 Cells

目的：探讨 Survivin表达降低时，对鼻咽癌细胞的增殖及凋亡的影响。方法运用 RNA 干扰技术，结合 RT-PCR，Western blot技术，观察干扰 Survivin效果。干扰 Survivn后用 MTT法和 TUNEL法检测鼻咽癌细胞增殖，凋亡的情况。用 Western-blot方法检测凋亡相关蛋白 PARP，Bcl-2和Bax的表达。结果 RT-PCR检测显示 Survivin-siRNA干扰组，Survivin mRNA表达明显下调，其相对表达量为0.26±0.02，抑制率为43.7％，与对照组相比差异具有统计学意义（P＜0.05）；Western blot检测显示 Survivin-siRNA组蛋白表达下调，其蛋白相对表达量下降了57％，与对照组相比差异具有统计学意义（P＜0.05）；MTT检测显示 Survivin-siRNA组对细胞增殖有明显抑制作用，在24，48和72 h的抑制率分别为21.9％，37.1％和29.6％，与对照组相比差异具有统计学意义（P＜0.05）；Tunel染色结果显示 Survivin-siRNA组细胞凋亡数明显增高。Western blot结果显示Survivin-siRNA组凋亡蛋白PARP（89KD）显著升高，为对照组的3.9倍，抑制凋亡蛋白Bcl-2降低，降低了70％，促进凋亡蛋白Bax升高，为对照组的2.4倍；同时也抑制AKT的磷酸化，磷酸化水平降低了57％，与对照组相比以上结果差异均具有统计学意义（P＜0.05）。结论干扰 Survivin表达，抑制鼻咽癌细胞增殖，促进细胞凋亡。Survivin基因可成为鼻咽癌基因治疗的一个潜在靶点。

Objective To explore the effect of decrease of Survivin expression on the proliferation and apoptosis of the naso-pharyngeal carcinoma cells.Methods To observe the effect of decrease of Survivin expression on nasopharyngeal carcinoma cell morphology using RNAi technology combined with RT-PCR and Western blot.Cell proliferation and apoptosis were de-tected by MTT and TUNEL assay.The expression of PARP,Bcl-2 and Bax was detected by Western blot.Results RT-PCR result showed the survivin mRNA expression in Surviving-siRNA groups was downregulated （about 0.26±0.02）and the inhibition ratio was 43.7% （P〈0.05）.MTT result showed cell proliferation rates were significantly different between 24,48 and 72 h after transfection.The cell inhibition rates were 21.9%,37.1% and 29.6%,respectively （P〈0.05）.West-ern blot result showed the Survivin protein expression in Survivin-siRNA2 groups was downregalated and the relative ex-pression level was reduced by 57% （P〈0.05）.TUNEL assay showed that cell apoptosis rate was increased obviously in surviving-siRNA groups.The expression of PARP （89KD）and Bax wasupregulated （3.9 and 2.4 fold change）and the ex-pression of Bcl-2 was downregulated （0.3 fold change）.The phosphorylation of AKT was inhibited when Survivin was down-regulated （about reduced by 5 7%）.Conclusion Silencing Survivin gene by siRNA can inhibit the proliferation of Na-sopharyngeal carcinoma cells and induce apoptosis.Survivin may become a potential gene for therapy target of nasopharynge-al carcinoma.