摘要
本文以食管癌Eca109、KYSE150细胞为研究对象,探讨铁死亡通路激活在雄黄抑制食管癌增殖和转移中的作用。采用CCK-8法检测抑制率及半数抑制浓度(IC_(50));克隆形成实验检测克隆形成能力;流式细胞检测细胞活性氧自由基(ROS)变化;透射电镜观察细胞超微结构;普鲁士蓝染色观察细胞内铁颗粒分布;免疫荧光染色检测细胞谷胱甘肽过氧化物酶4(GPX4)表达;划痕实验检测细胞迁移能力;Transwell小室实验检测细胞侵袭能力;Western blot检测细胞中E-cadherin、Slug、N-cadherin蛋白表达。结果显示,雄黄以时间及浓度依赖性方式抑制Eca109、KYSE150细胞增殖,作用Eca109、KYSE150细胞的IC_(50)分别为64.297、51.337μmol‧L^(-1);与空白组相比,雄黄2IC_(50)组Eca109、KYSE150细胞胞浆内大量线粒体发生肿胀并发现大小各异、多少不等的蓝染颗粒,且ROS荧光强度明显升高,GPX4蛋白表达明显降低(P<0.01);与雄黄组相比,雄黄+ZVAD-FMK组Eca109、KYSE150细胞增殖、迁移能力、穿膜数及Slug、N-cadherin蛋白表达明显升高,细胞抑制率及E-cadherin蛋白表达明显降低(P<0.05);雄黄+erastin组Eca109、KYSE150细胞增殖、迁移能力、穿膜数及Slug、N-cadherin蛋白表达明显降低,细胞抑制率及E-cadherin蛋白表达明显升高(P<0.05)。以上研究结果表明,雄黄可抑制Eca109、KYSE150细胞增殖,诱导细胞发生铁死亡现象,雄黄对食管癌细胞的增殖和转移作用可能通过部分铁死亡通路激活发挥作用。
This study was designed to investigate the role of activation of the ferroptosis pathway in the inhibition of esophageal cancer proliferation and metastasis by realgar,using esophageal cancer Eca109 and KYSE150 cells as the target cells.The rate of inhibition and half-inhibitory concentration(IC_(50))were measured by the CCK-8 method;clone formation ability was measured by clone formation assay;the changes in reactive oxygen species(ROS)were detected by flow cytometry;the ultrastructure of the cells was observed by transmission electron microscopy;the distribution of intracellular iron particles was observed by Prussian blue staining;the expression of glutathione peroxidase 4(GPX4)was detected by immunofluorescence staining;the scratch assay was used to detect the cell migration ability;the Transwell assay was used to detect the cell invasion ability;and western blotting was used to detect E-cadherin,Slug and N-cadherin protein expression in the cells.The results show that realgar inhibited the proliferation of Eca109 and KYSE150 cells in a time-and concentration-dependent manner,and the IC_(50) of Eca109 and KYSE150 cells was 64.297 and 51.337μmol‧L^(-1),respectively.Compared with the control group,many mitochondria in the cytoplasm of Eca109 and KYSE150 cells in the realgar 2IC_(50) group were swollen and blue-stained particles of different sizes and amounts were found,and ROS fluorescence intensity was significantly increased while GPX4 protein expression was significantly reduced(P<0.01).Compared with the realgar group,the proliferation,migration,membrane penetration and Slug and N-cadherin protein expression were significantly increased,and the cell inhibition rate and E-cadherin protein expression were significantly decreased in Eca109 and KYSE150 cells in the realgar+ZVAD-FMK group(P<0.05).The proliferation,migration,membrane penetration and Slug and N-cadherin protein expression were significantly decreased,and the cell inhibition rate and E-cadherin protein expression were significantly increased of E
作者
张雪莲
杨如意
王宏斌
孙兆
王君
王秋萍
陈发章
ZHANG Xue-lian;YANG Ru-yi;WANG Hong-bin;SUN Zhao;WANG Jun;WANG Qiu-ping;CHEN Fa-zhang(Medical College of Qinghai University,Xining 810001,China;Department of Integrative Medicine,Affiliated Hospital of Qinghai University,Xining 810001,China)
出处
《药学学报》
CAS
CSCD
北大核心
2022年第11期3322-3330,共9页
Acta Pharmaceutica Sinica
基金
青海省科学技术厅课题(2021-ZJ-730)。
关键词
食管癌
雄黄
铁死亡
细胞侵袭
细胞迁移
细胞增殖
esophageal cancer
realgar
ferroptosis
cell invasion
cell migration
cell proliferation
