缺氧诱导因子-1α和血管内皮生长因子干扰性RNA对人血管内皮细胞血管内皮生长因子表达的抑制  

Inhibition of expression of vascular endothelial growth factor by interfering RNA targeting hypoxia inducible factor-l~ and vascular endothelial growth factor in human vascular endothelial cells

作　　者：蒋剑[1] 夏晓波[1] 许惠卓[1] 熊思齐[1] 刘双珍[1] 李岩 

机构地区：[1]中南大学湘雅医院眼科,长沙410008 [2]University of Texas M. D. Anderson Cancer Center

出　　处：《中华眼底病杂志》2007年第6期422-425,共4页Chinese Journal of Ocular Fundus Diseases

基　　金：国家自然科学基金（030471853）

摘　　要：目的探讨缺氧诱导因子1α（HIF-1α）和血管内皮生长因子（VEGF）小片段干扰性RNA（siRNA）对人血管内皮细胞VEGF表达的影响。方法构建HIF-1α siRNA重组质粒。将体外培养的人血管内皮细胞（HUVEC12）分成常氧（20％）组和低氧（1％）组。低氧组中，采用脂质体Lipofectamine^TM 2000（LF2000）分别转染空载体质粒（空载体组）、HIF-1α siRNA（HIF-1α组）、VEGF165 siRNA（VEGF组）和HIF-1α siRNA＋VEGF165 siRNA（共转染组）。低氧组中未转染的细胞为对照组。采用逆转录-聚合酶链反应（RT—PCR）鉴定基因转染效率；RT—PCR和免疫细胞化学法检测各组细胞中VEGF基因和蛋白表达的变化。结果成功构建HIF-1α siRNA重组质粒。人血管内皮细胞转染24h后，HIF-1α siRNA和VEGF-1α siRNA重组质粒有表达。常氧组细胞中仅见微弱的VEGF mRNA和蛋白表达，而低氧组表达明显上调；HIF-1α组、VEGF组和共转染组表达较对照组减弱，其中共转染组抑制效果最明显。结论HIF-1α和VEGF-1α siRNA能有效抑制人血管内皮细胞VEGF的表达。Objective To investigate the effect of small interfering RNA（siRNA） targeting hypoxia inducible factor-1α （HIF-1α） and vascular endothelial growth factor （VEGF） on expression of VEGF in human vascular endothelial cells. Methods HIF-1α siRNA recombinant plasmid was constructed. Human vascular endothelial cells were cultured in vitro and divided into normoxia group （20% O2） and hypoxia group （1% O2）. Hypoxia group was then divided into control group, vector group, HIF-lα group （HIF-lα siRNA）, VEGF group （VEGF165 siRNA） and co transfection group （HIF lα siRNA＋VEGFL65 siRNA）. LipofectamineTM 2000 （LF2000） mediated vector plasmid was transfected to cells in each group except the control group. The expression of HIF-lα siRNA and VEGF165 siRNA recombinant plasmid were identified by reverse transcriptase polymerase chain reaction （RT-PCR）. The expression of VEGF mRNA and protein were detected by RT-PCR and immunocytochemical method. Results The expression of HIF-lα siRNA and VEGFI65 siRNA recombinant plasmid were detected 24 hours after transfected. The expression of VEGF mRNA and protein was faint in the normoxia group, but increased obviously in hypoxia group. The expression of VEGF mRNA and protein in the HIF-lα, VEGF and co-transfection groups were lower than which in the control group. Co-transfection group showed the highest inhibitory effect. Conclusion HIF-lα and VEGF165 siRNA can effectively inhibit the expression of VEGF in human vascular endothelial cells.

关 键 词：缺氧诱导因子1 Α亚基 血管内皮生长因子类 RNA干扰 内皮细胞 

分 类 号：R686[医药卫生—骨科学]