核转录因子-κB圈套策略对肝癌细胞HepG2凋亡的影响

Effects of Nuclear Factor-kappa B Decoy Oligonucleotide on the Apoptosis of HepG2 Cells

目的:观察NF-κB圈套寡核苷酸(Decoy ODNs)对NF-κB活性的影响,进一步探讨其对人肝癌细胞HepG2凋亡的作用机制。方法:设计NF-κB圈套寡核苷酸,将FITC标记的NF-κB Decoy ODNs转染HepG2,共聚焦显微镜观察其核内定位,描记生长曲线;凝胶阻滞法(EMSA)检测转染前后NF-κB活性;再分别用流式细胞术和TUNEL法检测细胞凋亡情况。Western blot检测HepG2细胞中Bcl-2和Fas的含量。结果:NF-κB Decoy ODNs转染HepG2细胞后定位于胞核,NF-κB活性明显下降,其显著抑制细胞生长,促进HepG2细胞凋亡;低活性NF-κB可以下调Bcl-2、增加Fas的表达水平。结论:NF-κB圈套寡核苷酸促进肝癌细胞HepG2凋亡的机制可能与其下调NF-κB的活性,使促凋亡蛋白Fas表达增加和降低抑凋蛋白Bcl-2表达有关。

Objective To investigate the effect of NF-KB decoy oligodeoxynucleotides （ODNs） on the activity of NF-κB and its mechanism in apoptosis of liver cancer cells HepG2. Methods The NF-κB decoy ODNs was designed, FITC-labeUed NF-κB decoy ODNs was transfected into HepG2, the nuclear localization was observed with confocal microscope, the growth curve was established;the activity change of NF-κB was detected with electrophoretic mobility shift assays （EMSA） pre- and post- transfection;the effect of low activity NF-κB on the cell proliferation was evaluated with flow cytometry and TUNEL, the expression change of Bcl-2 and Fas induced with low activity NF-κB were measured with Western blot. Results The decoy ODNs was located in the nuclei of HepG2 cells,the activity of NF-κB was significantly decreased,the growth of HepG2 was significantly inhibited,the apeptosis of liver cancer cells HepG2 was enhanced; the low activity of NF-κB upregulated the expression of Fas and downrcgulated the expression of Bcl-2. Conclusiorl The NF-κB decoy ODNs could increase the apoptosis of liver cancer ceils HepG2 and its mechanism may be related to the downregulation of NF-κB activity, upregulation of Fas expression and downregulation of Bcl-2 expression.