血管内皮生长因子反义RNA对EC9706人食管癌细胞抑制的作用

Inhibitory effect of vascular endothelial growth factor antisense RNA on the human esophageal cancer EC9706 cells

目的 观察血管内皮生长因子（VEGF）反义RNA对EC9706人食管癌细胞的抑制作用.方法 用脂质体法将反义VEGFcDNA质粒转染至EC9706人食管癌细胞,用噻唑蓝还原法（MTT法）检测EC9706细胞增殖,免疫组化SABC和逆转录-聚合酶链反应（RT-PCR）技术检测VEGF蛋白和VEGF mRNA表达水平.流式细胞术检测细胞凋亡和周期分布.并将转基因EC9706细胞接种于BALB/C裸鼠后肢皮下,4周后观察皮下成瘤情况.结果 被反义VEGFcDNA质粒转染的EC9706食管癌细胞有外源性VEGF反义基因的整合及表达,该细胞VEGF mRNA及蛋白的表达水平降低,但细胞生长增殖能力和增殖周期无明显改变,未发生明显凋亡现象 接种裸鼠28 d后,EC9706-wt组、EC9706-A组及EC9706-E组皮下移植瘤的潜伏期分别为（5.8±2.4）、（12.4±3.6）、（5.3±2.2）d,瘤体重量分别为（2.83 ±0.32）、（0.87±0.14）、（2.62 ±0.68）g,EC9706-A组与其他两组比较差异均有统计学意义（P＜0.05）.结论 VEGF反义RNA可抑制EC9706食管癌细胞VEGF表达和裸鼠体内肿瘤生长.

Objective To investigate the inhibitory effect of vascular endothelial growth factor （VEGF） antisense RNA on the growth of human esophageal cancer EC9706 cells in vitro. Methods The plasmid carrying VEGF antisense cDNA was transfected into EC9706 esophageal cancer cells, and the expression of VEGF was confirmed by reverse transcription-polymerase chain reaction （RT-PCR）. The expression level of VEGF mRNA and protein was examined in antisense group by RT-PCR and immunohistochemistry staining, respectively. The cell growth rate was detected by MTT assay. Apotosis and cell cycle in transfected cells were measured by flow cytometry. Results The expression of exogenous antisense VEGF mRNA was confirmed in transfected cells, and the VEGF protein and endogenous VEGF mRNA were significantly decreased. The growth rate of transfected cells was not inhibited. Apototic cells were not found in transfected cells. Twenty-eight days after inoculation in nude mice in EC9706-wt group, EC9706-A group and EC9706-E group, latency of subcutaneous tumors was （5.8 ± 2. 4 ）, （ 12. 4 ± 3.6） and （5.3 ± 2. 2） days, tumor weight was （2. 83 ± 0. 32 ）, （ 0. 87 ± 0. 14 ） and （ 2. 62 ± 0. 68 ） g, respectively. There was significant difference between EC9706-A group and other groups （ P 〈 0. 05）. Conclusion VEGF antisense RNA could decrease the expression of VEGF in esophageal cancer cells and cell proliferation in vivo, which may provide a useful theory basis for gene therapy of esophageal cancer.