Spautin-1通过抑制自噬可增强舒尼替尼诱导的肾癌细胞凋亡

Spautin-1 Enhances Sunitinib-induced Apoptosis in Renal Carcinoma Cells Via Inhibiting Autophagy

目的:研究spautin-1(一种自噬抑制剂)是否能抑制舒尼替尼诱导的肾癌细胞的自噬以及对舒尼替尼诱导的肾癌细胞凋亡的影响。方法:以肾癌细胞系786-O细胞为模型,Western Blot检测spautin-1对舒尼替尼诱导786-O细胞自噬的影响;Cell Counting Kit-8(CCK-8)检测spautin-1和舒尼替尼对786-O细胞增殖的影响;应用流式细胞术,检测spautin-1对舒尼替尼诱导的786-O细胞凋亡的影响;Western Blot检测spautin-1的促凋亡作用与PI3K/AKT/GSK3β信号通路及抗凋亡蛋白Bcl-2和Mcl-1的关系。结果:与舒尼替尼单独处理组相比,spautin-1能通过降低Beclin-1的表达显著抑制舒尼替尼在786-O细胞中诱导的自噬;Spautin-1和舒尼替尼联合作用明显增强舒尼替尼对786-O细胞增殖的抑制作用;Spautin-1能进一步增强舒尼替尼诱导的786-O细胞的凋亡;Spautin-1和舒尼替尼联合处理786-O细胞时,可以显著降低p-AKTSer473和p-GSK3βSer9的蛋白表达水平,增强GSK3β的活性,进而下调Bcl-2、Mcl-1的表达。结论:Spautin-1能通过抑制AKT活性并活化GSK3β,进一步降低抗凋亡蛋白Bcl-2、Mcl-1的表达,增强舒尼替尼诱导的肾癌细胞凋亡。

Objective: To investigate whether spautin-1, a specific and potent autophagy inhibitor, can enhance sunitinib-induced apoptosis in renal carcinoma cells via inhibiting autophagy. Methods: Renal carcinoma cell line 786-O was used as cellular model and Western Blot was used to detect the effect of spautin-1 on sunitinib-induced autophagy;Cell Counting Kit-8(CCK-8) was used to detect the effect of sunitinib on the proliferation of 786-O cells in the presence or absence of spautin-1. Flow cytometry was used to detect the effect of spautin-1 on sunitinib-induced apoptosis. Western Blot was used to detect the effects of spautin-1 on the PI3 K/AKT/GSK3β signaling pathway and the expression of the anti-apoptotic proteins Bcl-2 and Mcl-1. Results: Spautin-1 markedly inhibited sunitinib-induced autophagy by downregulating the autophagy protein beclin-1. Co-treatment with spautin-1 enhanced sunitinib-induced inhibition of cell proliferation. Spautin-1 enhanced sunitinib-induced apoptosis. Spautin-1 can decrease the phosphorylation levels of AKT and GSK3β in the PI3 K/AKT/GSK3β signaling pathway, co-treatment of spautin-1 and sunitinib downregulated anti-apoptotic proteins including Bcl-2 and Mcl-1. Conclusions: Spautin-1 enhanced sunitinib-induced autophagy and promoted sunitinib-induced cell apoptosis by activating GSK3β in PI3 K/AKT/GSK3β signaling pathway, which downregulated the expression of anti-apoptotic proteins Bcl-2 and Mcl-1.