siRNA 对人脐静脉内皮细胞中人剪切修复基因表达和凋亡的影响

摘要

Objective To study the effects on XPD gene expression and apoptosis of human umbilical vein endo-thelial cells (HUVECs) of transfecting small interfering RNA of xeroderma pigmentosum D.Methods Three specific re-combinant plasmids of XPD were constructed and identified.The silencing effects of XPDs in HUVCEs were assessed after siRNA transfection, and the siRNA with the best silence efficiency was selected among these 3 plasmids.Results West-ern blot and RT-PCR results showed that, compared with the control group, the transfection of all three recombinant plasmids of XPD-siRNAs significantly inhibited the expressions of XPD (P<0.01).Among these siRNAs, XPD-siR-NA-2 showed the best silence efficiency ( P<0.01 ) .Flow cytometry showed that, compared with control group, the XPD-siRNA-2 significantly reduced HUVCE apoptosis rate (P<0.01).Western blot results showed that, compared with control group, transfection of XPD-siRNA-2 significantly down-regulated the expression of Bax ( P<0.01 ) and up-regulated the expression of Bcl-2.Conclusion These findings prove that the expressions of XPD in HUVCEs can be effectively inhibited by the transfection of XPD-siRNA.The siRNA with the best silencing efficiency can inhibit the apoptosis of HUVEC, and Bax and Bcl-2 may take part in this process.%目的：向人脐静脉内皮细胞（HUVEC）中导入靶向人剪切修复基因D干扰序列（XPD-siRNA），观察其对HUVEC的XPD基因表达和凋亡的影响。方法以XPD为靶基因，构建3个特异性siRNA重组质粒，经转染后观察其对HUVEC中XPD的沉默作用；并转染沉默效率最佳的siRNA，观察其对细胞凋亡的影响。结果Western blot和RT-PCR结果显示，与阴性对照组相比，3个重组质粒XPD-siRNA的转染均能降低HUVEC中XPD的表达（P＜0.01），其中XPD-siRNA-2的沉默效率最高（P＜0.01）。流式细胞仪结果显示，与阴性对照组相比，XPD-siRNA-2组能降低HUVEC凋亡率（P＜0.01）。 Western blot结果显示，与阴性对照组相比，转染XPD-siRNA-2能下调Bax（P＜0.01）和上调Bcl-2（P＜0.01）的表达。结论 XPD-siRNA转染能有效沉默HUVEC中XPD基因的表达。所筛选沉默效率最佳的XPD-siRNA能抑制HUVEC凋亡，而Bax和Bcl-2可能参与了此过程。