补骨脂酚通过激活ROS强化TRAIL的抗肝细胞癌HepG2细胞作用的机制研究

目的:探究补骨脂酚(Bakuchiol,Bak)对肿瘤坏死因子相关凋亡诱导配体(Tumor necrosis factor-related apoptosis-inducing ligand,TRAIL)抗HepG2细胞作用的影响及内在机制。方法:常规培养HepG2细胞,给予梯度浓度的Bak处理,检测细胞活力。联合应用Bak与TRAIL处理,检测细胞活力。Western blot检测Bak处理后氧化应激水平、死亡受体4(Death Receptor 4,DR4)、DR5的表达变化。联合应用Bak与TRAIL检测凋亡情况。进而引入ROS清除剂NAC,联合NAC处理后,检测ROS、DR4、DR5以及凋亡情况。结果:Bak剂量依赖地抑制了HepG2细胞的活力,联合应用Bak+TRAIL对细胞活力的抑制作用优于单独用药。Bak处理后氧化应激水平升高,体现在ROS增加,GSH水平下降;Western blot检测发现Bak处理后DR4、DR5表达增加。联合应用Bak+TRAIL显著增加了细胞凋亡蛋白Bax的表达,抑制了抗凋亡蛋白Bcl2的表达。引入ROS阻断剂NAC处理后,与Bak+TRAIL组相比,ROS水平下降,DR4、DR5表达减少。凋亡检测发现NAC处理降低了Bak+TRAIL引起的细胞凋亡。结论:Bak可以显著增强TRAIL引起的HepG2细胞凋亡,该作用可能与Bak激活氧化应激进而上调DR4、DR5表达有关。

Bakuchiol Enhanced TRAIL Induced HepG2 Cell Apoptosis through Activating Oxidative Stress

ABSTRACT Objective: To investigate the roles of Bakuchiol (Bak) on HepG2 cell and the effects of co-treatment of Bak and Tumor necrosis factor- related apoptosis-inducing ligand (TRAIL) on HepG2 cells, as well as the inner mechanisms. Methods: Cells were treated with different concentrations of Bak or Bak+TRAILand cell vitality was detected 24 h later. After Bak treatment, cell oxidative stress levels were examined through DCF-DA staining and ROS, GSH detection. Western blot analysis was conducted to detected the expressions of Death receptor 4 (DR4) and DR5. After Bak+TRAIL treatment, cell apoptosis status was detected. Furthermore, ROS scavenger NAC was used to inhibit oxidative stress and DR4, DR5, Bax expressions were detected. Results: Both Bak and TRAIL can reduce HepG2 cell vitality. Co-treatment of Bak and TRAIL further reduced the cell vitality. Bak treatment increased ROS production, DR expressions and reduced cellular GSH level. Co-treatment of Bak and TRAIL induced higher level of apoptosis compared with single-drug treatment. Compared with the Bak + TRAIL group, NAC treatment reduced ROS level and reduced DR expression, as well as the Bak+TRAIL induced apoptosis levels. Conclusion: Bak enhances TRIAL induced HepG2 cell apoptosis and tumor growth via activating cellular oxidative stress and up regulating DR4 and DR5 expressions.