自噬在槲皮素诱导人膀胱癌BIU-87细胞凋亡中的作用

Role of autophagy in quercetin-induced apoptosis in human bladder carcinoma BIU-87 cells

目的研究自噬在槲皮素诱导的人膀胱癌BIU-87细胞凋亡中的作用。方法以不同浓度的槲皮素处理人膀胱癌BIU．87细胞，采用四甲基偶氮唑蓝（MTr）法检测槲皮素对BIU-87细胞增殖的抑制作用，采用单丹磺酰尸胺（MDC）荧光染色、Hochest33258染色分别观察槲皮素处理后BIU-87细胞的自噬和凋亡现象，采用流式细胞仪检测阻断自噬对BIU-87细胞凋亡的影响。以乳酸脱氢酶（LDH）检测试剂盒检测3一甲基腺嘌呤阻断自噬后对槲皮素细胞毒作用的影响。结果槲皮素对BIU-87细胞的增殖有明显的抑制作用，且此抑制作用呈明显的时间和浓度依赖性，200μmol／L槲皮素作用72h时对BIU．87细胞的增殖抑制率达89．2％。槲皮素可以诱导BIU-87细胞发生自噬和凋亡，并且自噬先于凋亡出现；阻断自噬后可使BIU-87细胞的凋亡峰明显增高。在槲皮素处理前后，分别以3．甲基腺嘌呤阻断自噬，可观察到槲皮素对BIU-87细胞的细胞毒作用均增强。结论槲皮素能明显抑制BIU-87细胞的增殖，并在诱导细胞凋亡之前先诱导自噬发生。在槲皮素诱导BIU-87细胞凋亡的过程中，自噬在早期能够保护BIU-87细胞，延迟凋亡，减少因槲皮素导致的细胞死亡。

Objective To explore the role of autophagy in quercetin （Que）-induced apoptosis in human bladder carcinoma BIU-87 cells in vitro. Methods To determine the proliferative inhibition by MTY colorimetric assay after treating BIU-87 cells with quercetin at various concentrations. To identify autophagy and apoptosis in the BIU-87 cells after Que treatment by monodansylcadaverin （MDC） and Hoechst 33258 fluorescent staining, respectively. To examine the cytotoxic effect of Que and influence of autophagy on apoptosis by studying LDH leakage rate and flow cytometry, after blocking the autophagy with 3- methlyadenine （3-MA）, a specific autophagy inhibitor. Results There was an obvious inhibitory effect of Que on the proliferation of BIU-87 cells in a time- and dose-dependent manner. The inhibition rate of BIU-87 cells after 200 μmol/L Que treatment for 72 hours was 89.2%. Autophagy and apoptosis were induced and detected in Que-treated BIU-87 cells and autophagy occurred earlier than apoptosis. The apoptosis peak became much higher after the autophagy was blocked. Whenever the autophagy was blocked before or after Que treatment, the Que-induced cytotoxicity in BIU-87 cells was enhanced. Conclusions Quercetin significantly inhibits the proliferation of BIU-87 cells, and the autophagy is induced earlier than apoptosis. In the process of Que-induced apoptosis of BIU-87 cells, autophagy may play a protective role at the initiation phase, delay apoptosis and reduce the Que-induced death of BIU-87 ceils.