Liu J Y, Li H, Liu Y, Shi C H, Liang Z L, Wang L H, Zhang Y, Zhao Y, Fan Y M, Wu B, Yu Y Z
Zhoukou Central Hospital, Zhoukou 466000, China.
Zhonghua Gan Zang Bing Za Zhi. 2019 Sep 20;27(9):693-697. doi: 10.3760/cma.j.issn.1007-3418.2019.09.007.
To investigate the effect and mechanism of XTP4 gene in apoptotic hepatoblastoma HepG2 cell line. HepG2 cells were transiently transfected with small interfering RNA of XTP4 genes, plasmid pcDNA3.1/myc-His(-) A-XTP4, and hepatitis B virus X protein transactivated x gene 4 (HBX protein trans-activate gene4, XTP4) and their respective negative controls. After 48h, the overexpression and interference expression condition of XTP4 in HepG2 cells were detected by Western blot. HepG2 cells apoptosis was detected by flow cytometry. The expression levels of apoptosis-related proteins P53, Bcl-2, Bax and Caspase-3 in HepG2 cells were detected by Western blot, and Bcl-2/Bax ratio was calculated. The chemiluminescence assay was used to detect activity of caspase-3 in HepG2 cells. The measured data were presented as ( ± s), and independent sample t-test was used for comparison between the two groups. HepG2 cells had successfully achieved the overexpression and interference expression of XTP4 protein. Compared with the control group, the overexpression of XTP4 in HepG2 cells had significantly decreased the number of apoptotic cells ( < 0.05), and increased Bcl-2/Bax ( < 0.05) ratio, but decreased the expression of P53 protein ( < 0.05). The protein expression of Caspase-3 and activity of caspase-3 was decreased ( < 0.05). However, interference with XTP4 expression in HepG2 cells had significantly increased the number of apoptotic cells ( < 0.05) and decreased Bcl-2/Bax ( < 0.05) ratio, but increased the expression of P53 protein ( < 0.05). The protein expression of Caspase-3 and activity of caspase-3 was increased (P<0.05). In HepG2 apoptosis XTP4 has inhibitory effect, and its effect on inhibiting HepG2 apoptosis may be achieved by regulating the Bcl-2/Bax ratio, and the P53 protein may be involved.
探讨XTP4基因对凋亡性肝癌HepG2细胞系的影响及其机制。将HepG2细胞分别用XTP4基因的小干扰RNA、质粒pcDNA3.1/myc-His(-) A-XTP4以及乙型肝炎病毒X蛋白反式激活基因4（HBX蛋白反式激活基因4，XTP4）及其各自的阴性对照进行瞬时转染。48小时后，通过蛋白质免疫印迹法检测HepG2细胞中XTP4的过表达和干扰表达情况。采用流式细胞术检测HepG2细胞凋亡情况。通过蛋白质免疫印迹法检测HepG2细胞中凋亡相关蛋白P53、Bcl-2、Bax和Caspase-3的表达水平，并计算Bcl-2/Bax比值。采用化学发光法检测HepG2细胞中Caspase-3的活性。测量数据以（±s）表示，两组间比较采用独立样本t检验。HepG2细胞成功实现了XTP4蛋白的过表达和干扰表达。与对照组相比，HepG2细胞中XTP4的过表达显著减少了凋亡细胞数量（P<0.05），增加了Bcl-2/Bax比值（P<0.05），但降低了P53蛋白的表达（P<0.05）。Caspase-3的蛋白表达和活性降低（P<0.05）。然而，干扰HepG2细胞中XTP4的表达显著增加了凋亡细胞数量（P<0.05），降低了Bcl-2/Bax比值（P<0.05），但增加了P53蛋白的表达（P<0.05）。Caspase-3的蛋白表达和活性增加（P<0.05）。在HepG2细胞凋亡中XTP4具有抑制作用，其抑制HepG2细胞凋亡的作用可能是通过调节Bcl-2/Bax比值实现的，P53蛋白可能参与其中。
