论文部分内容阅读
目的探讨反义VEGF165基因转染在抑制肿瘤血管生成和肿瘤生长中的作用。方法构建正义和反义VEGF165真核表达载体,用脂质体转染人神经母细胞瘤细胞SH-SY5Y,G418筛选稳定表达细胞克隆。应用RT-PCR证实外源性反义VEGFmRNA的表达,免疫细胞化学和ELISA检测VEGF蛋白的表达水平,MTT法检测肿瘤细胞体外生长情况,并接种于裸鼠皮下,观察体内肿瘤增殖能力。结果RT-PCR证实转染反义VEGF的细胞中有外源性反义VEGFmRNA的表达,免疫细胞化学和ELISA显示VEGF蛋白表达明显降低,MTT实验显示转染前后细胞体外生长速度基本一致,而转染反义VEGF的肿瘤细胞裸鼠体内生长速度明显减慢。结论反义VEGF基因转染能够有效抑制SH-SY5Y细胞内源性VEGF蛋白的表达,抑制裸鼠体内肿瘤的生长。
Objective To investigate the role of antisense VEGF165 gene transfection in inhibiting tumor angiogenesis and tumor growth. Methods The eukaryotic expression vector of sense and antisense VEGF165 was constructed and transfected into human neuroblastoma cells SH-SY5Y and G418 by Lipofectamine 2000 to screen stable clones. The expression of exogenous antisense VEGF mRNA was confirmed by RT-PCR. The expression of VEGF protein was detected by immunocytochemistry and ELISA. The in vitro growth of tumor cells was detected by MTT assay. The in vitro proliferation of tumor cells was inoculated in nude mice. Results RT-PCR confirmed the expression of exogenous antisense VEGF mRNA in antisense VEGF transfected cells. The immunocytochemistry and ELISA showed that the expression of VEGF protein was significantly decreased. The MTT assay showed that the growth rate of the transfected cells was basically the same, Tumor cells infected with antisense VEGF grew significantly slower in vivo. Conclusion Antisense VEGF gene transfection can effectively inhibit the expression of endogenous VEGF protein in SH-SY5Y cells and inhibit the growth of tumors in nude mice.