摘要目的 研究和厚朴酚(HNK)强化肿瘤坏死因子相关凋亡诱导配体(TRAIL)抗肝细胞癌HepG2细胞的作用及其机制.方法 常规培养HepG2细胞,首先给予梯度浓度HNK处理,观察其对肿瘤细胞活力的影响;Western blot法检测c-Jun氨基末端激酶(JNK)、死亡受体4(DR4)、 DR5表达水平变化.进而联合应用HNK与TRAIL,观察联合用药对肿瘤细胞活力的影响;检测凋亡相关蛋白表达水平明确凋亡情况.然后釆用JNK抑制剂SP600125阻断JNK通路,明确DR4、 DR5水平变化是否由JNK信号通路介导.最后构建裸鼠皮下肿瘤模型,明确在体内水平下,HNK对TRAIL的强化效果.结果 梯度浓度HNK处理细胞后细胞活力剂量依赖性降低(P＜ 0.05);联合应用TRAIL与HNK效果优于单纯用药(P ＜ 0.05);蛋白检测发现,HNK处理后pJNK水平上升,同时TRAIL受体DR4、DR5表达上调.联合应用HNK与TRAIL后,B淋巴细胞瘤因子2(bc12)显著下降而Bc12相关X蛋白(Bax)显著升高.SP600125阻断JNK通路后,与HNK + TRAIL组相比,DR4、DR5表达下降(P＜ 0.05),Bax表达下降而Bc12表达升高.在体内结果显示, TRAIL抑制了皮下肿瘤的生长,联合应用TRAIL与HNK抑制作用增强.结论 HNK可能通过激活JNK通路进而上调DR4、DR5表达来强化TRAIL对HepG2细胞的抑制作用.

abstractsObjective To investigate the effect and intrinsic mechanism of Honokiol (HNK) enhanced tumor necrosis factor-related apoptosis inducing ligand (TRAIL) on hepatocellular carcinoma HepG2 cells. Methods HepG2 cells were routinely cultured. Firstly, a concentration gradient of HNK was given to observe its effect on the vitality of tumor cells. Western blot were used to detect change in the expression levels of c-jun N-terminal kinase (JNK), death receptor 4 (DR4), and DR5. Secondly, we observed the effect of combined drugs (HNK and TRAIL) on the vitality of tumor cells. Apoptosis-related protein expression levels were detected to determine the apoptosis condition. Thirdly, JNK inhibitor SP600125 was used to block the JNK pathway, and it was evaluated whether JNK signaling pathway mediated the DR4 and DR5 levels and finally, the subcutaneous tumor model of nude mice was constructed, and enhancement effect of HNK on TRAIL was confirmed in vivo. Results Cell vitality was decreased (P ＜ 0.05) in a dose-dependent manner after treatment with gradient HNK. Combined effect of TRAIL and HNK was superior to that of single drug administration (P ＜ 0.05). Western blot analysis showed that pJNK level increased after HNK treatment and TRAIL receptor DR4 and DR5 expression were up-regulated. Combined application of HNK and TRAIL, B lymphocyte tumor factor 2 (BCL2) decreased significantly while Bc12 related X protein (Bax) increased significantly. Blocking JNK pathway by SP600125, the expression of DR4 and DR5 decreased (P ＜ 0.05), Bax expression decreased and Bc12 expression increased compared with HNK+TRAIL group. In vivo results showed that TRAIL inhibited the growth of subcutaneous tumors, and enhanced inhibition effect in combination with TRAIL and HNK. Conclusion HNK may enhance the inhibitory effect of TRAIL on HepG2 cells by activating JNK pathway and then upregulating the expression of DR4 and DR5.