TGF-β_1对VEGF在膀胱肿瘤血管生成中的调节作用

Down-regulation Effect of TGF-β_1 on Expression of VEGF in Bladder Cancer Cell Line

目的采用RNA干扰(RNAi)技术沉默转化生长因子基因(TGF-β1),观察RNAi的基因沉默效应及对人膀胱癌细胞株(EJ细胞)中血管内皮生长因子基因(VEGF)表达的影响,探讨TGF-β1对VEGF在膀胱肿瘤血管生成中的调节作用。方法构建基因TGF-β1特异性siRNA表达载体,Real time-PCR及ELISA筛选出抑制效率最高的靶序列,分成EJ细胞组、对照组(TGF-β1组)、重组质粒组(TGF-β1siRNA表达载体组),转染EJ细胞后,Real time-PCR检测VEGF mRNA的水平。结果成功构建基因TGF-β1特异性siRNA表达载体,Real time-PCR及ELISA筛选出最佳抑制效果的序列〔TGF-β1mRNA表达水平为0.92±0.19;其细胞培养上清液中TGF-β1蛋白表达水平为(50.08±5.85)pg/mL〕转染后,各组的VEGF mRNA表达水平以重组质粒组最低(P<0.05);VEGF mRNA的表达随TGF-β1基因沉默而减少(P<0.05)。结论抑制TGF-β1基因能下调VEGF基因的表达,TGF-β1基因可能通过诱导VEGF基因的表达实现对膀胱肿瘤血管生成的调控。

Objective To explore the down-regulation effect of transforming growth factor-β1 （TGF-β1） on the expression of vascular endothelial growth factor （VEGF） in bladder cancer cell line EJ cell strain. Methods The siRNA expression vectors of TGF-β1 gene were constructed, the highest inhibition target sequence was screened and selected by Real time-PCR and ELISA, and then the vectors were transfected into EJ cells, the expression level of VEGF mRNA was detected by Real time-PCR. Results TGF-β1 targeting expression vectors were successfully constructed, Real time-PCR and ELISA screened the highest inhibition target sequence （the lowest expression level of TGF-β1 mRNA was 0. 92±0. 19; ELISA result was 50. 08±5. 85）. The relative expression level of VEGF mRNA in TGF-β1 siRNA group was the lowest, indicating that the expression of VEGF was decreased by silencing TGF-β1 gene. Conclusion Inhibition of TGF-β1 gene expression could down regulate the expression of VEGF in bladder neoplasm.