血管内皮生长因子-C shRNA对肝癌HepG2细胞增殖及侵袭能力的影响

目的 检测靶向血管内皮生长因子-C(vasenlar endothelial growth factor-C,VEGF-C)shRNA质粒载体对肝癌HepG2细胞增殖及侵袭能力的影响.方法 构建VEGF-C shRNA质粒载体,脂质体转染方法转入肝癌HepG2细胞.通过倒置荧光显微镜及流式细胞仪检测细胞的转染率;RT-PCR及Western blot检测转染细胞内VEGF-C mRNA及蛋白的表达变化;MTT法检测细胞增殖抑制率;人工基底膜体外侵袭实验检测细胞侵袭能力.结果 VEGF-C shRNA稳定转染后,肝癌HepG2细胞内VEGF-C mRNA及蛋白表达水平显著下降;VEGF-C shRNA对肝癌HepG2细胞具有明显的增殖抑制作用,其抑制增殖效应呈时间依赖性;VEGF-C shRNA可有效抑制肝癌HepG2细胞的人工基底膜体外侵袭能力,抑制率为51.54%.结论 VEGF-C在肝癌增殖、侵袭转移中发挥重要作用;通过RNA干扰技术实现VEGF-C沉默,在肝癌的基因治疗中具有较好的应用前景.

Effects of short hairpin RNA expression plasmid targeting vascular endothelial growth factor-C on the proliferation and invasion of HepG2 cells

Objective To investigate the effects of short hairpin RNA (shRNA) expression plasmid targeting vascular endothelial growth factor-C (VEGF-C) on the proliferation and invasion of HepG2 cells. Methods The VEGF-C shRNA plasmid vector labeled with green fluorescent protein was constructed and stably transfected into HepG2 cells. The transfected cells were sorted by G418 and visualized by fluorescent microscope and assayed by flow cytometry. Expression of VEGF-C in transfected cells was determined by RT-PCR and Western blot. The inhibition rates of the cell proliferation and invasion were determined by MTT assay and reconstituted basement membrane invasion assay, respectively. Results VEGF-C shRNA effectively downregulated VEGF-C mRNA and protein expression in HepG2 cells, and it also effectively inhibited the proliferation of HepG2 cells in a time-dependent manner. The invasion capacity of HepG2 cells was inhibited by VEGF-C shRNA, and the inhibition rate was 51.54%. Conclusions VEGF-C plays an important role in tumor proliferation, invasion and metastasis. RNA interfering technology that targets VEGF-C may serve as a potential therapeutic intervention in the treatment of human hepatic cancer.