AMPK-mediated autophagy inhibits apoptosis in cisplatin-treated tumour cells.
AMPK-mediated autophagy inhibits apoptosis in cisplatin-treated tumour cells.
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DOI：
10.1111/j.1582-4934.2009.00663.x
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发表时间：
2009-09
影响因子：
5.3
通讯作者：
Trajkovic V
中科院分区：
文献类型：
--
作者：
Harhaji-Trajkovic L;Vilimanovich U;Kravic-Stevovic T;Bumbasirevic V;Trajkovic V
利用人U251胶质瘤、大鼠C6胶质瘤和小鼠L929纤维肉瘤细胞系研究了自噬在顺铂抗癌作用中的作用。顺铂治疗后，在肿瘤细胞中观察到自噬的剂量和时间依赖性诱导，如自噬诱导蛋白beclin-1的上调和随后出现的吖啶橙染色的酸性自噬囊泡所示。顺铂处理的细胞中的自噬体的存在也证实了电子显微镜。用溶酶体抑制剂巴弗洛霉素A1和氯喹或PI 3激酶抑制剂渥曼青霉素抑制自噬，显著增强顺铂触发的氧化应激和半胱天冬酶激活，导致DNA片段化和凋亡性细胞死亡增加。自噬的保护作用的机制显然涉及干扰顺铂诱导的Bcl-2家族蛋白的调节，因为自噬的抑制增强了顺铂介导的促凋亡Bax的上调和抗凋亡Bcl-2的下调。顺铂处理的细胞中的自噬诱导之前，腺苷一磷酸激活的蛋白激酶（AMPK）的激活和伴随的下调哺乳动物雷帕霉素靶蛋白（mTOR）介导的p70 S6激酶磷酸化。顺铂触发自噬的能力被小干扰RNA（siRNA）介导的AMPK沉默降低，而用mTOR siRNA转染足以触发肿瘤细胞中的自噬。最后，siRNA介导的AMPK下调和AMPK抑制剂化合物C增加顺铂诱导的肿瘤细胞死亡，而mTOR siRNA和AMPK激活剂二甲双胍保护肿瘤细胞免受顺铂的影响。总之，这些数据表明，顺铂触发的AMPK激活和随后的mTOR活性抑制可以诱导自噬反应，保护肿瘤细胞免受顺铂介导的凋亡性死亡。
The role of autophagy in cisplatin anticancer action was investigated using human U251 glioma, rat C6 glioma and mouse L929 fibrosarcoma cell lines. A dose- and time-dependent induction of autophagy was observed in tumour cells following cisplatin treatment, as demonstrated by up-regulation of autophagy-inducing protein beclin-1 and subsequent appearance of acridine orange-stained acidic autophagic vesicles. The presence of autophagosomes in cisplatin-treated cells was also confirmed by electron microscopy. Inhibition of autophagy with lysosomal inhibitors bafilomycin A1 and chloroquine, or a PI3 kinase inhibitor wortmannin, markedly augmented cisplatin-triggered oxidative stress and caspase activation, leading to an increase in DNA fragmentation and apoptotic cell death. The mechanisms underlying the protective effect of autophagy apparently involved the interference with cisplatin-induced modulation of Bcl-2 family proteins, as inhibition of autophagy potentiated cisplatin-mediated up-regulation of proapoptotic Bax and down-regulation of anti-apoptotic Bcl-2. Autophagy induction in cisplatin-treated cells was preceded by activation of adenosine monophosphate-activated protein kinase (AMPK) and concomitant down-regulation of mammalian target of rapamycin (mTOR)-mediated phosphorylation of p70S6 kinase. The ability of cisplatin to trigger autophagy was reduced by small interfering RNA (siRNA)-mediated AMPK silencing, while transfection with mTOR siRNA was sufficient to trigger autophagy in tumour cells. Finally, siRNA-mediated AMPK down-regulation and AMPK inhibitor compound C increased cisplatin-induced tumour cell death, while mTOR siRNA and AMPK activator metformin protected tumour cells from cisplatin. Taken together, these data suggest that cisplatin-triggered activation of AMPK and subsequent suppression of mTOR activity can induce an autophagic response that protects tumour cells from cisplatin-mediated apoptotic death.