Survivin-siRNA对裸鼠视网膜母细胞瘤移植瘤的抑制研究  

The Experiment Study on Survivin-siRNA Inhibiting the Growth of Y79 Retinoblastoma Implantation Model in Nude Mice Vitreous Cavity

作　　者：吴荒[1] 孙莹[2] 张小猛[1] 刘喜春[3] 张灵[3] 吴雅臻[1] 

机构地区：[1]吉林大学第二医院,吉林长春130041 [2]吉林大学第一医院 [3]吉林大学前列腺疾病防治研究中心

出　　处：《中国实验诊断学》2010年第2期179-182,共4页Chinese Journal of Laboratory Diagnosis

摘　　要：目的构建Survivin-siRNA真核重组质粒,转染裸鼠视网膜母细胞瘤玻璃体腔移植瘤中,探讨其对肿瘤组织中Survivin表达的抑制作用及诱导凋亡情况。方法构建针对Survivin mRNA特异的靶序列的Survivin-siRNA以及对照Scramble-siRNA。将Y79细胞种植至21只裸鼠玻璃体腔内,9d后随机平均分为mock对照组(注射K-PBS);Scramble对照组(注射带有非同源的Scramble-siRNA重组质粒);Survivin实验组(注射特异性Survivin-siRNA重组质粒)。10d后摘除肿瘤组织,利用半定量RT-PCR检测肿瘤组织Survivin mRNA;采用Western blot方法分析Survivin蛋白;HE染色、Survivin及Ki67免疫组化染色;TUNEL法检测肿瘤组织凋亡。结果真核重组质粒及视网膜母细胞瘤裸鼠玻璃体腔移植瘤模型均构建成功。半定量RT-PCR显示Survivin-siRNA在mRNA水平上抑制了Survivin基因转录(P<0.01),Scramble组与mock组无差别(P>0.05)。Western blot显示与mock组和Scramble组相比,实验组细胞的Survivin蛋白表达显著降低(P<0.01)。HE染色普通光镜下观察,实验组与两对照组相比,在瘤组织内可见到较多死亡细胞;实验组移植瘤组织中Survivin蛋白表达与mock组和scramble组相比明显下降;实验组Ki67的表达较对照组下降明显,增殖指数明显下降(P<0.01)。TUNEL法显示实验组肿瘤组织内可见大量细胞核染为棕黄色的凋亡细胞,对照组可见少许凋亡细胞,实验组凋亡指数明显高于对照组(P<0.01)。结论成功构建的Survivin-siRNA重组质粒可以抑制裸鼠视网膜母细胞瘤移植瘤中Survivin基因转录和蛋白表达,诱导了肿瘤细胞凋亡。Objective To construct survivin siRNA eukaryotic expression vectors and evaluate their effect on the growth of Y79 retinoblastoma implantation model in nude mice vitreous cavity. Methods Survivin-siRNA and Scramble-siRNA were constructed. Y79 cells were injected into vitreous cavity of twenty-one nude mice. Mice were divided equally into three groups （mock, Scramble- siRNA, Survivin-siRNA） nine days later. Tumors were enucleated after ten days. Semi-quantitive R-T-PCR was used to detect Survivin mRNA in tumor tissues. Western Blotting was used to analyze the expression of Survivin protein. The tumor tissues were observed by HE staining. The expression of Survivin and Ki67 were observed by immunohistochemical staining. TUNEL method was used to analyze the apoptosis of retinoblastoma cells. Results Both eukaryotic expression vectors and retinoblastoma implantation model in nude mice vitreous cavity were established successfully. The expression of Survivin was significantly inhibited at mRNA level comparing with the control groups（ P 〈 0.01 ）, and no.significant difference was found between two control groups（ P 〉 0.05） by semi-quantitive RT- PCR. Comparing with the control groups,the expression of Survivin was significantly inhibited at protein level by Western Blotting（ P 〈 0.01）,and no significant difference was found between two control groups（ P 〉 0.05）. More dead cells were detected in experimental group by HE staining. The expression of Sttrvivin and Ki67 was inhibited significantly（ P 〈 0.01 ）in experimental group. More apoptosis Cells were ＇observed in experimental group, and apoptosis index was significant higher in experimental group than in control groups（ P 〈 0.01 ）by TUNEL. Conclusion Survivin-siRNA can be used for interrupting the gene transcription and downregulating the protein expression for Y79 retinoblastoma in nude mice vitreous cavity. Survivin-siRNA induces the apoptosis of retinoblastoma cells.

关 键 词：RNA干扰 SURVIVIN 凋亡 视网膜母细胞瘤 

分 类 号：R739.7[医药卫生—肿瘤]