摘要
目的探讨小檗碱对前列腺癌细胞铁死亡的影响及其作用机制。方法体外培养前列腺癌DU145和PC-3细胞,采用CCK-8检测不同浓度(1.560、3.125、6.250、12.500、25.000、50.000、100.000μmol/L)小檗碱对前列腺癌细胞增殖活力的影响,并计算半数抑制浓度(half-inhibitory concentration,IC50);采用谷胱甘肽(glutathione,GSH)试剂盒检测细胞内GSH水平;胱氨酸定量分析试剂盒检测细胞内胱氨酸水平,DCFH-DA荧光探针法检测细胞内活性氧(reactive oxygen species,ROS)水平;采用荧光探针法检测细胞内的Fe^(2+)水平;q RT-PCR检测铁死亡相关基因(GPX4、COX2、FTH1和SLC7A11)的表达水平。结果各浓度小檗碱均可抑制DU145和PC-3细胞增殖活力(均P<0.05),且呈浓度依赖性,IC50分别为19.94μmol/L和20.18μmol/L。铁死亡抑制剂Ferrostatin1(Fer-1)可逆转小檗碱(20μmol/L)对DU145和PC-3细胞增殖活力的抑制作用(均P<0.001),但细胞凋亡和焦亡抑制剂不能恢复细胞活力。20μmol/L小檗碱可明显降低DU145和PC-3细胞内GSH水平和胱氨酸水平(均P<0.001),上调脂质ROS水平和Fe^(2+)水平(均P<0.001),但Fer-1可逆转小檗碱诱导的细胞内脂质ROS水平和Fe^(2+)水平(均P<0.001),并恢复GSH和胱氨酸水平(均P<0.001)。此外,小檗碱显著下调DU145和PC-3细胞中SLC7A11和GPX4的表达水平(均P<0.05),并上调COX2和FTH1的表达水平(均P<0.01)。过表达SLC7A11可逆转小檗碱对前列腺癌细胞铁死亡的促进作用(均P<0.001)。结论小檗碱可能通过抑制SLC7A11表达促进前列腺癌细胞内脂质ROS累积和Fe^(2+)水平上调,进而导致细胞发生铁死亡。
Objective To investigate the effect of berberine on ferroptosis in prostate cancer cells and its mechanism.Methods Prostate cancer DU145 and PC⁃3 cells were cultured in vitro,and the effects of berberine with different concentrations(1.560,3.125,6.250,12.500,25.000,50.000,100.000μmol/L)on the proliferation ability of prostate cancer cells were evaluated by CCK⁃8 assay,and the half⁃inhibitory concentration(IC50)were calculated.The levels of glutathione(GSH),cysteine,reactive oxygen species(ROS),and Fe^(2+)in both DU145 and PC⁃3 cells were determined by GSH detection assay kit,cysteine assay kit,DCFH⁃DA fluorescent probe assay,and the fluorescent probe,respectively.qRT⁃PCR was performed to detect the expression levels of ferroptosis⁃related genes(GPX4,COX2,FTH1 and SLC7A11).Results Berberine at various concentrations inhibited the proliferation abilities of DU145 and PC⁃3 cells in a concentration⁃dependent way(all P<0.05),with IC50 values of 19.94μmol/L and 20.18μmol/L,respectively.The ferroptosis inhibitor Ferrostatin1(Fer⁃1)reversed the inhibitory effect of berberine(20μmol/L)on cell proliferation in DU145 and PC⁃3 cells(all P<0.001),whereas cell apoptosis and pyroptosis inhibitors could not restore cell viability.Treatment with berberine at the concentration of 20μmol/L significantly decreased the levels of GSH and cysteine in DU145 and PC⁃3 cells(all P<0.001),and increased the levels of lipid ROS and Fe^(2+)(all P<0.001).However,the Fer⁃1 could reverse berberine⁃induced intracellular lipid ROS level and Fe^(2+)level(all P<0.001),and restored the levels of GSH and cysteine(all P<0.001).In addition,berberine significantly down⁃regulated the expression levels of SLC7A11 and GPX4 in DU145 and PC⁃3 cells(all P<0.05),while up⁃regulated the expression levels of COX2 and FTH1(all P<0.01).Overexpression of SLC7A11 reversed the promotive effect of berberine on ferroptosis in prostate cancer cells(all P<0.001).Conclusions Berberine promotes intracellular lipid ROS accumulation and up
作者
穆克飞
叶纪伟
沈远径
张赛
郭新武
MU Kefei;YE Jiwei;SHEN Yuanjing;ZHANG Sai;GUO Xinwu(Department of Urology and Andrology,Nanyang Second General Hospital,Nanyang 473000,China)
出处
《中国癌症防治杂志》
CAS
2023年第3期278-284,共7页
CHINESE JOURNAL OF ONCOLOGY PREVENTION AND TREATMENT
基金
南阳市科技发展计划项目(KJGG2018124)。
关键词
前列腺癌
小檗碱
铁死亡
脂质氧化
Prostate cancer
Berberine
Ferroptosis
Lipid oxidation
