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TGF-β1对VEGF在膀胱肿瘤血管生成中的调节作用
Down-regulation Effect of TGF-β1 on Expression of VEGF in Bladder Cancer Cell Line
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摘要: 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-β1 siRNA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-β1 mRNA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 （PVEGF mRNA的表达随TGF-β1基因沉默而减少（PTGF-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。Abstract: 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-β1mRNA 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.
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