谷胱甘肽过氧化物酶4介导的铁死亡参与阿霉素对乳腺癌细胞抑制作用研究

GPX4-mediated ferroptosis is involved in the inhibitory effect of adriamycin on breast cancer cells

目的:探讨谷胱甘肽过氧化物酶4(GPX4)介导的铁死亡是否参与阿霉素对乳腺癌的抑制过程,并探讨其增加阿霉素敏感性的可能机制。方法:通过实时荧光定量逆转录聚合酶链反应(qRT-PCR)实验和免疫蛋白印迹法(Western Blot),检测正常乳腺细胞MCF10A、HCC1937细胞和乳腺癌细胞MCF-7、MDA-MB-231细胞中GPX4的表达水平;通过慢病毒转染技术构建稳定敲低GPX4的乳腺癌细胞MCF-7,并通过qRT-PCR和Western Blot实验检测敲低效率;通过CCK8实验检测阿霉素对乳腺癌细胞的细胞毒性;通过谷胱甘肽(GSH)检测试剂盒、丙二醛(MDA)检测试剂盒、铁离子(Fe^(2+))浓度检测试剂盒检测敲低GPX4后乳腺癌细胞的铁死亡水平。结果:乳腺癌细胞中GPX4的蛋白水平及mRNA水平高于正常乳腺细胞(P<0.05);与对照组细胞相比,敲低GPX4表达的乳腺癌细胞中GSH水平降低(P<0.05),MDA和Fe^(2+)水平升高(P<0.05),阿霉素作用后细胞增殖能力下降(P<0.05),而铁死亡抑制剂ferrostatin-1可以逆转GPX4敲低后阿霉素对乳腺癌细胞增殖能力的抑制作用(P<0.05)。结论:沉默GPX4可以促进乳腺癌细胞铁死亡过程,最终促进乳腺癌对阿霉素的敏感性。

Objective:To investigate the effect of GPX4-mediated iron death on adriamycin sensitivity in breast cancer and its related molecular mechanism.Methods:qRT-PCR and western blot assay were used to explore the expression of GPX4 in the normal breast cells MCF10A and HCC1937 and the breast cancer cells MCF-7 and MDA-MB-231.Lentivirus transfection technique was used to construct MCF-7 breast cancer cell with stable knockdown of GPX4,and the knockdown efficiency was detected by qRT-PCR and western blot assay.CCK8 assay was used to detect the sensitivity of breast cancer cells to adriamycin.The GSH concentration,MDA level and iron ion level of breast cancer cells after GPX4 knockdown were detected by GSH detection kit,MDA detection kit and iron ion concentration detection kit.Results:The expression of GPX4 in breast cancer cells was significantly higher than those in normal breast cells.Knockdown of GPX4 significantly reduced the level of GSH,promoted the levels of MDA and Fe^(2+),and significantly enhanced the sensitivity of GPX4 to adriamycin in breast cancer cells,while ferrostatin 1 reversed the sensitivity of GPX4 to adriamycin in breast cancer cells.Conclusion:Silencing GPX4 can promote the ferroptosis of breast cancer cells,and ultimately promote the sensitivity of breast cancer to adriamycin.