【摘要】 背景与目的:利用小干扰RNA(small interfering RNA,siRNA)抑制哺乳动物基因表达已成为研究基因功能的一种有效方法。本研究探讨人胰岛素样生长因子1类受体(insulin-like growth factor 1 receptor,IGF1R)的短发夹环RNA对人肝癌细胞株SMMC7721裸鼠移植瘤的抑制作用。方法:设计并合成特异性靶向IGF1R的siRNA片段,构建SMMC7721-IGF1R-siRNA表达质粒,将其转入SMMC7721细胞。通过G418筛选出稳定株,移植裸鼠成瘤。同时设立对照组。根据体积的均数值绘制肿瘤生长曲线,以HE染色法观察质粒处理后瘤组织的病理改变,Westernblot检测肿瘤组织中IGF1R的表达变化,免疫组化SP法检测检测瘤组织内微血管密度,原位末端标记法(TUNEL法)检测肿瘤细胞的凋亡情况。结果:SMMC7721-IGF1R-siRNA组肿瘤体积与SMMC7721-IGF1R-mutation组、SMMC7721组相比差异有统计学意义(P<0.05)。病理学检查及TUNEL检测发现,SMMC7721-IGF1R-siRNA组肿瘤生长受到抑制,细胞凋亡指数[(50.2±6.4)%]明显高于SMMC7721-IGF1R-mutation组[(5.4±1.0)%]和SMMC7721组[(6.0±2.1)%](P<0.05)。SMMC7721-IGF1R-siRNA组与SMMC7721-IGF1R-mutation组、SMMC7721相比,瘤内IGFIR蛋白表达明显下调。SMMC7721-IGF1R-siRNA组微血管密度(11.3±4.4)与SMMC7721-IGF1R-mutation组(36.7±7.6)、SMMC7721组(28.4±6.5)相比明显下降(P<0.05)。结论:构建的SMMC7721-IGF1R-siRNA具有RNA干扰作用,能抑制SMMC7721细胞裸鼠移植瘤的生长。更多还原

【Abstract】 BACKGROUND & OBJECTIVE: Using small interfering RNA (siRNA) to inhibit mammal gene expression becomes an effective technique in studying gene function. This study was to investigate the effect of insulin-like growth factor 1 receptor (IGF1R) siRNA on the growth of human liver cancer SMMC7721 cell xenograft in nude mice. METHODS: siRNA targeting IGF1R was designed,and plasmid SMMC7721-IGF1R-siRNA was constructed and transfected into SMMC7721 cells (SMMC7721-IGF1R-siRNA cells); the cells transfected with SMMC7721-IGF1R-mutation (SMMC7721-IGF1R-mutation cells) were used as negative control,and untransfected cells as empty control. Stable cell clones were screened by G418,and transplanted into nude mice to establish cancer xenograft. Tumor growth was monitored. Tumor morphology was observed with HE staining. The expression of IGF1R protein in tumor tissues was detected by Western blot. Microvessel density (MVD) in tumor tissues was detected by SP immunohistochemistry. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. RESULTS: The tumor volume was significantly smaller in SMMC7721-IGF1R-siRNA group than in SMMC7721-IGF1R-mutation group and SMMC7721 group (P < 0.05). Necrosis and cell apoptosis were found in SMMC7721-IGF1R-siRNA group. The expression of IGF1R protein was significantly lower in SMMC7721-IGF1R-siRNA group than in SMMC7721-IGF1R-mutation group and SMMC7721 group (P < 0.05). MVD was significantly lower in SMMC7721-IGF1R-siRNA group than in SMMC7721-IGF1R-mutation group and SMMC7721 group (11.3±4.4 vs. 36.7±7.6 and 28.4±6.5,P < 0.05). The apoptosis rate of tumor cells was significantly higher in SMMC7721-IGF1R-siRNA group than in SMMC7721-IGF1R-mutation group and SMMC7721 group (50.2±6.4)% vs. (5.4±1.0)% or (6.0±2.1)%,P < 0.05. CONCLUSION: IGF1R siRNA can inhibit the growth of SMMC7721 cell xenograft in nude mice.更多还原