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目的:研究PI3K通路在蛋白酶体抑制剂MG132诱导卵巢癌细胞自噬中的作用。方法:MG132处理多种卵巢癌细胞(SKOV3、OVCAR3、A2870)后,光镜观察胞质内囊泡的形成,吖啶橙(AO)染色观察酸性囊泡的形成,Western blot检测自噬特异性蛋白LC3的变化。PI3K抑制剂(渥曼青霉素与3-MA)联合MG132处理多种卵巢癌细胞后,AO染色及West-ern blot检测自噬的变化。采用shRNA技术下调OVCAR3细胞的Beclin 1表达,MG132处理细胞,AO染色观察酸性囊泡的形成。结果:在多种卵巢癌细胞系中,MG132可诱导酸性囊泡形成及LC3-Ⅰ向LC3-Ⅱ的转化增强,PI3K抑制剂与MG132联合应用与单独应用MG132相比自噬水平无改变。下调卵巢癌细胞中Beclin 1表达后,MG132诱导的囊泡形成无改变。结论:MG132通过PI3K非依赖性途径诱导多种卵巢癌细胞发生自噬。
AIM: To investigate the role of PI3K pathway in autophagy induced by proteasome inhibitor MG132 in ovarian cancer cells. Methods: MG132 cells were treated with various ovarian cancer cells (SKOV3, OVCAR3, A2870). The formation of intracytoplasmic vesicles was observed under light microscope. The formation of acid vesicles was observed by acridine orange (AO) staining and the autophagy specificity Changes in protein LC3. PI3K inhibitor (wortmannin and 3-MA) combined with MG132 treatment of a variety of ovarian cancer cells, AO staining and West-ern blot detection of autophagy changes. Beclin 1 expression was down-regulated by shRNA in OVCAR3 cells, MG132 cells were treated with AO staining to observe the formation of acid vesicles. RESULTS: In many ovarian cancer cell lines, MG132 induced the formation of acidic vesicles and the enhanced LC3-I conversion to LC3-Ⅱ. The combination of PI3K inhibitor and MG132 did not change the level of autophagy compared with MG132 alone. After down-regulating Beclin 1 expression in ovarian cancer cells, MG132-induced vesicle formation did not change. Conclusion: MG132 induces autophagy in a variety of ovarian cancer cells through a PI3K-independent pathway.