靶向血管内皮生长因子C短发夹式RNA对人乳腺癌MCF-7细胞生物学特性的影响

目的 研究重组质粒表达载体介导的短发夹式RNA(shRNA)对乳腺癌MCF-7细胞株中血管内皮生长因子C(VEGF-C)的表达及乳腺癌细胞增殖、侵袭能力的影响.方法 脂质体介导重组质粒pSIREN-VEGF-C转染乳腺癌MCF-7细胞株,嘌呤酶素筛选阳性克隆;荧光定量PCR及Western blot检测转染前后乳腺癌MCF-7细胞中VEGF-C基因的表达;MTT法及Transwell体外侵袭实验检测转染前后乳腺癌MCF-7细胞的增殖及侵袭能力.结果 转染重组质粒后乳腺癌MCF-7细胞中VEGF-C mRNA及蛋白表达均明显下调,抑制率分别为95%及100%(P<0.05);MTT法和 Transwell体外侵袭实验显示,与阴性质粒组和空白对照组比较,转染重组质粒后各时间段乳腺癌MCF-7细胞增殖均明显受抑制(P<0.05),同时穿膜的肿瘤细胞数也明显减少(29.0±1.9)个与(59.0±2.1)个和(61.0±2.2)个相比,P<0.05].结论 表达VEGF-C shRNA的重组质粒载体pSIREN-VEGF-C在乳腺癌MCF-7细胞中能高效、特异地发挥靶基因沉默作用,并对乳腺癌细胞的增殖及侵袭能力有显著抑制作用.

Role of short haipin RNA targeting vascular endothelial growth factor C on biological characteristics of human breast cancer cell MCF-7

Objective To study the effect of short haipin RNA ( shRNA) on expression of vascular endothelial growth factor C ( VEGF-C) and proliferation and invasion behavior of human breast cancer cell MCF-7.Methods The recombinant vector (pSIREN-VEGF-C) was transfected into the human breast cancer cell MCF-7 by liposome and the positive transfected cell clones were screened with puromycin.Expression of VEGF-C in MCF-7 cells after gene transfer was detected by real-time quantitative PCR and Western blot assay,respectively.Proliferation and invasion ability of transfected cells were analyzed by MTT and Transwell filter.Results The expressions of VEGF-C mRNA and protein were decreased markedly compared with the control group after the transfection and the inhibitive ratio was 95% and 100% respectively (P<0.05).The proliferation of MCF-7 cells transfected by pSIREN-VEGF-C,measured with MTT assays,was significantly decended (P < 0.05).The invasion ability of passing through the Transwell filter of MCF-7 cells transfected by pSIREN-VEGF-C were declined evidently (P <0.05).Conclusion The recombinant vector (pSIREN-VEGF-C) have been proved not only to be effective and specific for down-regulation of VEGF-C,but also can inhibit the proliferation and invasion of MCF-7 cells significantly.