抑制自噬晚期阶段增强榭皮素诱导的胶质瘤细胞凋亡

摘要

目的 研究抑制自噬不同阶段对榭皮素作用于胶质瘤细胞的影响,并对其机制进行初步探讨.方法 通过3-甲基腺嘌呤(3-MA)及转染Beclin 1 shRNA质粒抑制自噬早期阶段,通过氯喹(CQ)抑制自噬晚期阶段.噻唑蓝比色法(MTT)检测细胞活性.通过透射电镜及Western Blot检测自噬水平.通过流式细胞技术及Western Blot检测细胞凋亡.结果 榭皮素可以诱导胶质瘤细胞凋亡及自噬的发生.抑制自噬的早期阶段会减弱榭皮素对胶质瘤细胞的毒性作用,抑制自噬的晚期阶段可以增强榭皮素诱导的胶质瘤细胞凋亡.结论 氯喹与榭皮素联合用药具有协同作用,可能对胶质瘤的治疗提供新的思路.%Objective The cytotoxicity of Quercetin (Quer) to glioma when inhibiting autophagy at different stages was explored and the preliminary mechanism was discussed.Methods 3-methyladenine (3-MA) and Beclin 1 shRNA were used to inhibit the early stage of autophagy.Chloroquine (CQ) was used to block the late stage of autophagy.Methylthiazolyldiphenyl-tetrazolium bromide assay (MTF) was adopted to assess the cell viability induced by Quer when autophagy was inhibited at different stages.Transmission electron microscopy and Western Blot were used to detect autophagy level.Flow-cytometry and Western Blot were used to measure cell apoptosis.Results Cell viability was inhibited and autophagy was induced by Quer in a dose-dependent manner.Inhibition of autophagy at an early stage attenuated Quer-induced cytotoxicity.In contrast,blockade of autophagy flux at late stage by CQ enhanced the cytotoxicity of Quer,and caused accumulation of degradative autophagic vacuoles and robust apoptosis.Conclusion CQ and Quer have a synergistic effect,which may provide a novel opportunity for glioma therapy.