Cyclin G1对肝癌细胞HepG2放射敏感度的影响及其机制

Effect of Cyclin G1 on Radiosensitivity of Hepatocellular Carcinoma HepG2 Cells and Its Mechanism

目的探讨Cyclin G1对肝癌HepG2细胞放射敏感度的影响及其可能的分子机制。方法通过转染Cyclin G1-siRNA构建HepG2-Cyclin G1-siRNA细胞,通过成克隆分析观察Cyclin G1对肝癌细胞放射敏感度的影响,流式细胞仪检测肝癌细胞细胞周期分布,ELISA法检测乏氧因子HIF-1α及ROS的表达,Western blot检测凋亡相关蛋白Bcl-2、Bax的表达。结果 q-PCR显示HepG2-Cyclin G1-siRNA细胞中,Cyclin G1表达下降至正常细胞的43.5%左右,成克隆分析显示Cyclin G1-siRNA可使HepG2细胞放射敏感度增高,与正常对照组相比,放射增敏比SER_(Dq)为1.41。流式细胞分析显示HepG2-Cyclin G1-siRNA细胞与正常HepG2细胞相比,G_2/M期细胞比例增加,G_0/G_1期细胞比例下降;ELISA显示X线照射可引起HepG2细胞的HIF-1α升高,而Cyclin G1-siRNA可以引起HIF-1α的含量显著下降。ROS含量在X射线照射后表现为轻度升高,Cyclin G1-siRNA可以使ROS含量增加,联合X线照射变化更明显。Western blot显示X线照射及Cyclin G1-siRNA均可使BCL-2表达下降及Bax表达增加,其中HepG2联合X线照射组改变最明显。结论 Cyclin G1-siRNA可以上调肝癌细胞HepG2的放射敏感度,其机制可能与调节HIF-1α、ROS及凋亡相关蛋白有关。

Objective To investigate the effect of Cyclin G1 on the radiosensitivity of hepatocellular carcinoma HepG2 cells and its possible mechanism.Methods HepG2-Cyclin G1-siRNA cells was constructed through the transfection of Cyclin G1-siRNA to HepG2 cells,and the effect of Cyclin G1 on radiosensitivity of HepG2 cells was observed.Flow cytometry was used to monitor the change of cell cycle.The expression of hypoxia-inducible factor-1α（HIF-1α） and ROS were detected by ELISA analysis,and Western blot was used to detect the expression of Bcl-2 and Bax.Results q-PCR showed the expression of Cyclin G1 in HepG2-Cyclin G1-siRNA cells was decreased to 43.5% of normal control group.Clone formation analysis showed that Cyclin G1-siRNA could increase the radiosensitivity of HepG2 cells,SERDq=1.41,compared with normal control group.Flow cytometry analysis showed that the cells in G2/M phase were increased and the cells in G0/G1 phase were decreased in HepG2-Cyclin G1-siRNA cells.ELISA analysis showed that the HIF-1α of HepG2 cells could be induced by X-ray irradiation,while Cyclin G1-siRNA could cause a significant decrease in the content of HIF-1α.ROS content in X-ray-irradiated cells showed mild increase,with no significant difference.Cyclin G1-siRNA could increase the content of ROS,moreover,the change was more significant when it combined with X-ray radiation.Western blot showed that X-ray irradiation and Cyclin G1-siRNA could decrease the expression of BCL-2 and increase the expression of Bax,and the changes of HepG2-Cyclin G1-siRNA combined with X-ray irradiation group were the most obvious.Conclusion Cyclin G1-siRNA could upregulate the radiosensitivity of hepatocellular carcinoma cell line HepG2,and its mechanism may be related to the regulation of HIF-1,ROS and apoptosis-related proteins expressions.