siRNA腺病毒体内抑制Survivin基因表达诱导裸鼠肿瘤细胞凋亡的研究

目的利用腺病毒siRNA载体介导的RNA干扰技术,在免疫缺陷鼠肝癌肿瘤动物模型中通过靶向沉默细胞凋亡因子Survivin的表达,探讨其在活体动物肿瘤组织中的作用。方法利用肝癌细胞株HepG2构建裸鼠肝癌肿瘤细胞动物模型,并按照病毒注射剂量分组,将已构建的Survivin-siRNA重组腺病毒注射入裸鼠肿瘤内,观察肿瘤生长状况并绘制肿瘤生长曲线,利用TUNEL反应测定肿瘤组织细胞内DNA的片段化观察细胞凋亡。结果注射AdsiR- NA-Survivin高、低剂量组肿瘤生长明显受到抑制,且抑制程度与注射的腺病毒浓度呈正相关;TUNEL实验表明注射AdsiRNA-Survivin高、低剂量组的肿瘤组织标本可见大量细胞核呈黄褐色、核质浓缩、细胞形态不规则的凋亡细胞。高剂量组、低剂量组、AdsiRNA-U6组、PBS组凋亡细胞数依次减少。结论腺病毒siRNA载体系统可应用于活体动物试验中;AdsiRNA-Survivin对裸鼠人肝癌移植瘤有明显抑制作用,与肿瘤感染数或浓度成正比,且能促进人肝癌肿瘤细胞的凋亡。

Tumor cells apoptosis in nude mice induced by inhibited Survivin gene expression by adenovirus-mediated siRNA

Objective In order to research the Survivin gene's function on the animal tumor,we inhibited the expression of the Survivin in the nude mice hepatocarcinoma animal model via adenovirus-mediated siRNA system.Methods The hepatocarcinoma nude mice model by the hepatocarcinoma cells HepG2 was constructed,divided into four groups accord- ing to the injection dose of the AdsiRNA-Survivin.The constructed Survivin-siRNA adenovirus was injected into the hep- atocarcinoma nude mice tumor,and the tumor's growth was observed,and the tumor cells apoptosis was detected by the TUNEL kit which can assay sliced DNA in tumor cells.Results The growth of the tumors injected with high or low dose of AdsiRNA-Survivin were obviously inhibited,and the level of inhibition was in positive correlation with injected dose of adenovirus.The results of the TUNEL test showed that a lot of the apoptosis cells which turned to he tawny, with concentrated nucleoid and irregular cell shape could be observed in the high or low injection dose of tumors.The number of the apoptosis cells was decreased in turn by the groups as high dose group,low dose group,AdsiRNA-U6 group and PBS group.Conclusion Our results demonstrated that the adenovirus mediated siRNA system could be used for animal experiments in vivo,and the AdsiRNA-Survivin could efficiently inhibit the tumor growth and induce the tumor cells apoptosis,whose effect is dose-depended.