RNA干扰抑制结肠癌细胞血管内皮生长因子表达

目的 应用RNA干扰技术抑制结肠癌血管内皮生长因子（VEGF）表达。方法 将VEGF基因作为RNA干扰的靶区，通过E-RNAi网上提供的服务，设计两个特异的RNA干扰序列，将其装入含U6启动子的载体上，构建成抗VEGF基因的小发夹样RNA（shRNA）表达载体，再转染人结肠癌细胞HT29，通过RT-PCR、Northern杂交、免疫荧光和Western杂交，观察VEGF表达受抑的程度。结果 成功构建了两种抗VEGF基因的shRNA表达载体，RT-PCR、Northern杂交、免疫荧光和Western杂交，均发现其能明显抑制HT29细胞VEGF基因的表达，抑制率分别达42%、88%、73%和82%。结论 针对VEGF基因的shRNA表达载体能够明显抑制结肠癌细胞VEGF基因的表达。

Inhibition of Vascular Endothelial Growth Factor by RNA Interference in HT29 Cells

Objective:To inhibit the expression of Vascular Endothelial Growth Factor (VEGF) by RNA interference in HT29 cells. Methods:Using the service of E-RNAi, we designed and constructed two kinds of shRNA expression vectors which were aimed at the VEGF gene, then transfected them into colon cancer HT29 cells by lipofectamineTM 2000. The level of VEGF mRNA was determined by RT-PCR and Northern blotting. The protein expression of VEGF was examined by Immunofluoresence staining and Western blotting. Results:The two kinds of VEGF specific shRNA expression were found to efficiently inhibit the expression of VEGF in HT29 cells, the inhibition rate showed by RT-PCR, Northern blotting, Immunofluoresence staining and Western vectors blotting were 42%, 88%,73% and 82%, respectively. Conclusion:The expression of VEGF could be inhibited by RNA interference in HT29 cells.