VEGFsiRNA对人舌癌Tca8113细胞移植瘤血管生成影响的研究

The influence of VEGFsiRNA on the angiogenesis of human tongue squamous cell carcinoma(Tca8113) xenografts in vivo.

目的:观察小干扰RNA(small interfering RNA,siRNA)抑制血管内皮生长因子(vascular endothelial growthfactor,VEGF)表达对裸鼠皮下人舌癌Tca8113细胞移植瘤血管生成的影响。方法:构建人舌癌Tca8113细胞20只裸鼠皮下移植瘤模型,随机分为4组。将两对针对VEGF的siRNA真核表达载体(Pu-VEGF-siRNA1,Pu-VEGF-siRNA2)脂质体法作瘤体内及瘤周注射;以注射空质粒组作实验对照组;未注射组作空白对照组。注射1次/3d×10次,末次注射3d后,剥离瘤体,RT-PCR检测瘤组织VEGF-mRNA表达,免疫印迹技术检测瘤组织VEGF蛋白表达。免疫组化染色法观察VEGF阳性染色及微血管参数。结果:Pu-VEGF-siRNA2组移植瘤VEGF-mRNA表达及VEGF蛋白表达、总体微血管密度、有腔微血管密度、有腔血管平均血管周长及管腔面积均较空质粒组及空白对照组低(p<0.05)。而Pu-VEGF-siRNA1组未观察到上述差别(p>0.05)。结论:siRNA能在体内抑制舌癌VEGF表达,有效减少肿瘤血管生成。不同的干扰片段体内作用效果存在差异。

Objective：To investigate the influence of VEGFsiRNA on the angiogenesis of human tongue squamous cell carcinoma （Tca8113） xenografts in vivo, Method-Palpable human tongue cancer tumors were established on the backs of nude mice by subcutaneous injection of cultured cells （Tca8113）, The animals were divided into 4 groups randomly （n= 5/group）. Two siRNA targeting VEGF constructed in eukaryotic expression vector （PU-VEGF-siRNAI,PU-VEGF- siRNA2） were injected introtumor and peritumor with lipofectamine2000, respectively. Tumors injected vector without siRNA and Non-injected tumors were used as experimental and negative controls. Animals were injected one time every 3 days for total 10 times. Three days after the last injection, the tumors were excised, the expression of VEGF in xenograft tumors were detected by reverse transcription polymerse chain reaction （RT-PCR） and immunohistochemistry.Microvessel density, opening microvessel density,mean perimeter and lumen area of microvessel in xenograft tumors were detected by immunohistochemical stain. Result：Compared to the experimental and negative controls, the expression of VEGF and microvessel values were descreased in the PU-VEGF-siRNA2 group （p 〈 0.05）. But there were not statistically significantly difference of those between PU-VEGF-siRNAI group and two controls （p 〉 0.05）. Conclusion： VEGF-siRNA can reduce VEGF expression in vivo, inhibit agiogenesis of tumor. Different VEGF-siRNA may lead to different effect in vivo.