Ku80高表达对肝癌SMMC7721细胞凋亡的影响

目的 观察Ku80高表达对SMMC7721肝癌细胞凋亡的影响。方法 将Ku80基因转染到SMMC7721细胞;流式细胞术和TUNEL法检测细胞凋亡;Western blot检测凋亡相关蛋白表达。结果 筛选获得稳定高表达Ku80的克隆细胞;流式检测显示,Ku80高表达克隆的凋亡率(9.44±1.52)％和(9.26±1.72)％与对照组(1.81±0.15)％和(1.83±0.25％)比较轻度增高,差异有统计学意义(P ＜0.05);TUNEL实验显示,Ku80高表达克隆形成裸鼠皮下瘤的凋亡率(9.3±2.0)％和(10.0±2.1)％与对照组(3.5±1.0)％和(3.6±1.1)％比较轻度增高,差异有统计学意义(P＜0.05);Western blot检测显示,Ku80高表达克隆cleaved PARP-1、active Caspase-3和cleaved Caspase-9的表达与对照组比较明显增高,bcl-2的表达减低,而bax无变化。结论 体内外实验均证实Ku80高表达可引起SMMC7721细胞凋亡轻度增加。

Effect of Ku80 re-expression on cell apoptosis in hepatocellular carcinoma SMMC7721 cells

Objective To investigate the effect of Ku80 re-expression on cell apoptosis in hepatocellular carcinoma (HCC) SMMC7721 cells. Methods PcDNA3. 1 ( + )-myc-his-Ku80 and pcDNA3. 1( + )-myc-his expressive plasmids were transfected into Ku80 deficient SMMC7721 HCC cells. The cell apoptotic rates were analyzed by facial action coding system (FACS). The expression of cell apoptosis related regulators were analyzed by Western blotting. Cell apoptotic levels in xenograft tumor tissue were detected by TUNEL assay. Results The Western blotting analysis confirmed that Ku80-transfected cells expressed high protein levels of Ku80,whereas the vector-transfected and the parental SMMC7721 cells lacked Ku80 expression. FACS analysis indicated that, the apoptotic rates in SMMC7721, vector-transfected cells and the two Ku80-expressing clones were (9.44 ±1.52)％, (9.26 ±1.72)％ ; (1.81 ±0. 15)％ and (1.83 ±0. 25) ％, respectively. There was a significant difference in cell apoptotic rate between Ku80-expressing clones group and SMMC7721 or vector-transfected cells group ( P ＜ 0. 05 ). TUNEL assay also showed that,the cell apoptotic rates in tumor tissue derived from SMMC7721, vector-transfected cells and the two Ku80-expressing clones were (9. 3 ± 2. 0) ％, ( 10. 0 ± 2. 1 ) ％ ; (3.5 ± 1.0) ％ and (3.6 ± 1.1 ) ％, respectively. There was a significant difference in cell apoptotic rate between Ku80-expressing clones group and SMMC7721 or Vector-transfected cells group (P ＜0. 05). Western blotting further confirmed that levels of cleaved PARP-1, active Caspase-3 and cleaved Caspase-9 significantly increased in the Ku80-expressing clones compared with vector-transfected or parental SMMC7721 cells,whereas the expression levels of bcl-2 decreased. The expression level of bax remained unchanged. Conclusion Ku80 re-expression slightly induces cell apoptosis in SMMC7721 cells in vitro and in vivo.