槲皮素通过GAS5调控GSK-3β/β-catenin逆转乳腺癌阿霉素耐药

Quercetin reverses adriamycin resistance by activating GSK-3β/β-catenin pathways through GAS5 in breast cancer cells

目的:探究槲皮素逆转阿霉素耐药作用及其可能机制。方法:采取CCK-8法检测槲皮素(ADR)对阿霉素耐药细胞的细胞毒性及对阿霉素耐药的逆转效果;平板克隆形成实验检测细胞增殖;Hoechst染色检测细胞凋亡;流式细胞仪检测细胞内罗丹明123(Rh123)积累变化;qRT-PCR法检测细胞中GAS5、ABCB1 RNA表达水平;Western blot检测GSK-3β、β-catenin、c-MYC、cyclin D1、ABCB1蛋白表达水平。结果:槲皮素(10、20、40μmol/L)可以明显增强MCF-7/ADR对ADR的敏感性,抑制细胞的增殖,增加细胞内Rh123积累,促进GAS5、GSK-3β的表达,抑制β-catenin、c-MYC、cyclin D1、ABCB1的表达;细胞转染si-GAS5后,GAS5、GSK-3β表达明显下降,β-catenin、c-MYC、cyclin D1、ABCB1表达明显升高;当加入槲皮素(40μmol/L)与si-GAS5共处理后,与sncRNA加槲皮素共处理对照组相比,这些蛋白的表达都发生了逆转性的回复作用。结论:槲皮素可以通过GAS5介导GSK-3β/β-catenin通路抑制ABCB1的表达从而逆转乳腺癌细胞阿霉素耐药。

AIM: To investigate the ability of quercetin to reverse acquire adriamycin(ADR) resistance and explored its probably mechanism. METHODS: The CCK-8 assay was used to detect the cytotoxicity of quercetin in MCF-7/ADR cells and the reversal effect of ADR. The colony formation assay and Hoechst 332582 staining were used to detect the cell proliferation, cell apoptosis and the accumulation of rhodamine 123(Rh123) respectively. The RNA expression levels of GAS5 and ABCB1 were detected by qRT-PCR. The protein expression levels of GSK-3β, β-catenin, c-MYC, cyclin D1, and ABCB1 were detected by Western blot. RESULTS: Quercetin(10, 20, 40 μmol/L) significantly enhanced the sensitivity of MCF-7/ADR to ADR, inhibitd cell proliferation, and increased the intracellular accumulation of Rh123. Treatment with quercetin in MCF-7/ADR cells, the expression levels of GAS5 and GSK-3β were increased, whereas the expression levels of β-catenin, c-myc, cyclin D1 and ABCB1 were decreased. Further research revealed that reduction of GAS5 by RNA interference(si-GAS5) induced inhibitory effects on the expressions of GAS5 and GSK-3β, and enhanced the expressions of β-catenin, c-myc, cyclin D1, and ABCB1. Furthermore, treatment by quercetin combined with si-GAS5 in MCF-7/ADR cells, the expressions of these proteins were effectively reversed in comparison to quercetin combined with siRNA negative control(sncRNA). CONCLUSION: Quercetin increases the expression of GAS5 by GSK-3β/β-catenin signaling pathway, which inhibits the expression of ABCB1, ultimately reversing ADR resistance in the MCF-7/ADR cells.