千金藤素通过调控PI3K/AKT/mTOR信号通路诱导卵巢癌SKOV3细胞自噬

目的:研究千金藤素(cepharanthine)对人卵巢癌SKOV3细胞自噬的诱导作用,并探讨其可能的作用机制。方法:体外培养SKOV3细胞,CCK-8法检测cepharanthine(0、2、4、8、16和32μmol/L)对SKOV3细胞活力的影响;吖啶橙染色观察细胞酸性自噬泡;Western blot法检测cepharanthine作用后自噬相关蛋白LC3及PI3K/AKT/mTOR信号通路蛋白表达的变化。结果:Cepharanthine抑制卵巢癌SKOV3细胞活力,呈一定的剂量依赖关系(P<0.05)。Cepharanthine能显著增加SKOV3细胞内酸性自噬泡数量。Western blot结果表明在8和16μmol/L的cepharanthine作用下,卵巢癌SKOV3细胞中LC3-Ⅱ蛋白表达水平显著升高(P<0.01)。信号通路蛋白研究表明,cepharanthine显著降低p-AKT和p-mTOR的蛋白水平(P<0.01),AKT和mTOR总蛋白水平均无明显变化。自噬抑制剂3-甲基腺嘌呤可增强cepharanthine对SKOV3细胞活力的抑制作用(P<0.01)。结论:Cepharanthine可诱导人卵巢癌SKOV3细胞自噬,其作用机制可能与抑制PI3K/AKT/mTOR信号通路有关。

Cepharanthine induces autophagy via PI3K/AKT/mTOR signaling pathway in ovarian cancer SKOV3 cells

AIM:To investigate the autophagy of human ovarian cancer SKOV3 cells induced by cepharanthine and to explore its mechanism.METHODS:The effect of cepharanthine on the viability of ovarian cancer SKOV3 cells was measured by CCK-8 assay.The SKOV3 cells were treated with cepharanthine,and then the formation of autophagosome was observed with acridine orange staining under fluorescence microscope.The protein levels of LC3,AKT,p-AKT,mTOR,p-mTOR and GAPDH in the SKOV3 cells treated with cepharanthine were determined by Western blot.RESULTS:Cepharanthine significantly inhibited the viability of ovarian cancer SKOV3 cells in a dose-dependent manner(P<0.05).The number of the intracellular acidic autophagosomes with bright red fluorescence was significantly increased after cepharanthine treatment in the SKOV3 cells.The expression of LC3-II in SKOV3 cells was significantly enhanced after cepharanthine treatment.Furthermore,treatment with cepharanthine in the SKOV3 cells also resulted in a significant down-regulation of phosphorylated form of AKT and mTOR(P<0.01),while the total protein level was not changed.Combination of cepharanthine and 3-methyladenine resulted in a substantial decrease in the cell viability compared with using cepharanthine alone.CONCLUSION:Cepharanthine significantly inhibits the growth of human ovarian cancer SKOV3 cells and induces the autophagy,which may be correlated with down-regulation of PI3K/AKT/mTOR signaling pathway.