NVP-BEZ235抑制肝癌细胞HepG2的增殖和迁移及机制研究

目的 探究NVP-BEZ235对肝癌细胞HepG2增殖和迁移的影响及其机制。方法 NVP-BEZ235(0、25、50、100、250、500 nmol·L^-1)处理HepG2细胞48 h后,采用MTT法检测细胞的活性;通过形态学观察和Hoechst 33342染色法,检测NVP-BEZ235对HepG2细胞增殖、凋亡的影响;采用划痕实验、Transwell迁移实验,检测NVP-BEZ235对HepG2细胞迁移能力的影响;免疫印迹法检测PI3K/Akt/mTOR信号通路、上皮-间质转化(EMT)以及Six1/Ezrin信号轴等标志物的表达情况;采用免疫荧光实验检测EMT相关标志物的表达情况。结果 NVP-BEZ235可明显抑制HepG2细胞的增殖,且呈浓度依赖性;NVP-BEZ235可诱导HepG2细胞发生凋亡;NVP-BEZ235可抑制HepG2细胞的迁移能力。在NVP-BEZ235的作用下,p-Akt^Ser473 、p-S6^Thr389 、p-4E-BP1^Thr37/46 的表达下调,上皮标志物E-cadherin表达上调,间质标志物Vimentin、Snail表达下调,Six1和p-Ezrin^Tyr353 表达下调。结论 NVP-BEZ235可有效抑制HepG2细胞的增殖和迁移,其机制可能与NVP-BEZ235降低p-Akt^Ser473 、p-S6^Thr389 、p-4E-BP1^Thr37/46 的表达,抑制Six1/Ezrin信号轴及逆转EMT的进程有关。

Inhibitory effect of NVP-BEZ235 on proliferation and migration of hepatocellular carcinoma cell HepG2 and its mechanism

Aim To evaluate the efficacy of NVP-BEZ235 on the proliferation and migration of HepG2 human hepatocellular carcinoma cell in vitro ,and to investigate the molecular mechanism.Methods The cytotoxicity of NVP-BEZ235 on HepG2 cells treated with NVP-BEZ235(0,25,50,100,250,500 nmol·L^-1) was detected by MTT assay.The ability of NVP-BEZ235 to modulate HepG2 cells proliferation and apoptosis was detected by morphology assay and Hoechst 33342 staining.The motility of NVP-BEZ235 treatment on HepG2 cells was determined by wound healing assay and Transwell assay.The expression levels of the related protein including PI3K/Akt/mTOR signal pathway,epithelial-mesenchymal transition(EMT) process and Six1/Ezrin signal axis were detected by Western blot.The expression levels of EMT markers were also detected by immunofluorescence staining.Results The cell viability significantly decreased in HepG2 cells treated with NVP-BEZ235 compared with control group,which showed a dose-dependent manner.NVP-BEZ235 could induce apoptosis and inhibit HepG2 cell migration.E-cadherin protein expression significantly increased;however,the expressions of p-Akt^Ser473 ,p-S6^Thr389 ,p-4E-BP1^Thr37/46 ,Vimentin,Snail,Six1 and p-Ezrin^Tyr353 decreased in NVP-BEZ235 treated HepG2 cells compared with those in control group.Conclusions NVP-BEZ235 could effectively suppress the proliferation and migration of HepG2 cells,which may be related to the down-regulation of p-Akt^Ser473 ,p-S6^Thr389 ,p-4E-BP1^Thr37/46 and modulating both Six1/Ezrin signal axis and the EMT process by NVP-BEZ235 treatment.