荜茇酰胺导致胃癌细胞铁死亡的作用研究  

Study on the mechanism of piperlongumine in iron death of gastric cancer cells

作　　者：张学松[1] 宋毓飞[1] 康锦钰 黄璐捷 ZHANG Xue-song;SONG Yu-fei;KANG Jin-yu;HUANG Lu-jie(Department of Gastroenterology,Ningbo Medical Center Li Huili Hospital,Ningbo 315040,Zhejiang Province,China;Department of General Practice,Ningbo Medical Center Li Huili Hospital,Ningbo 315040,Zhejiang Province,China;Medical College of Ningbo University,Ningbo 315040,Zhejiang Province,China)

机构地区：[1]宁波市医疗中心李惠利医院消化内科,浙江宁波315040 [2]宁波市医疗中心李惠利医院全科医学,浙江宁波315040 [3]宁波大学医学院,浙江宁波315040

出　　处：《中国临床药理学杂志》2020年第10期1280-1283,共4页The Chinese Journal of Clinical Pharmacology

基　　金：浙江省基础公益研究计划基金资助项目(LGF19H030008)。

摘　　要：目的研究荜茇酰胺(piperlongumine,PIP)诱导胃癌细胞HGC-27铁死亡的作用。方法采用梯度浓度的PIP干预HGC-27细胞,细胞随机分为对照组和PIP组。对照组为常规培养的细胞,PIP组分别以终浓度为5,10,20μmol·L^-1的PIP进行干预。以细胞计数试剂盒-8(CCK-8)检测细胞活力的改变,以二氯荧光素(DCFH-DA)探针检测活性氧(ROS)的产生,以蛋白质印迹法检测谷胱甘肽过氧化物酶4(GPX4)、胱氨酸-谷氨酸反向转运体亚基xCT蛋白、金属离子转运体1(DMT1)的表达水平,以试剂盒检测细胞内丙二醛(MDA)含量。用ROS抑制剂乙酰半胱氨酸(NAC)预处理,将HGC-27细胞随机分为对照组,PIP组,NAC+PIP组。对照组为常规培养的细胞,NAC+PIP组先用5μmol·L^-1的NAC预处理12 h,而PIP组和NAC+PIP组中PIP均为10μmol·L^-1的终浓度进行干预。同样检测细胞活力,ROS产生及相关蛋白表达。结果PIP干预后,细胞活力显著下调,具有剂量依赖性,最高以20μmol·L^-1的PIP干预后,细胞活性可降至(50.34±0.13)%。探针检测显示细胞内ROS的水平增高,细胞内MDA水平增高,同样具有剂量依赖性,最高以20μmol·L^-1的PIP干预后,MDA水平可升至(15.41±0.17)μmol·mL^-1。同时铁死亡关键蛋白GPX4、xCT的表达下调而DMT1的表达水平上调。NAC预处理后可以显著抑制铁死亡,细胞活力增高,24 h后NAC+PIP组细胞活性为(65.42±0.41)%,PIP组细胞活性为(58.21±0.56)%,差异有统计学意义(P<0.05)。同时ROS的产生下调,细胞内MDA含量下调,铁死亡关键蛋白GPX4、DMT1的表达上调而xCT的表达水平下调,其中NAC+PIP组MDA水平为(6.24±0.21)μmol·mL^-1,PIP组MDA水平为(11.13±0.03)μmol·mL^-1,差异有统计学意义(P<0.05)。结论荜茇酰胺可以通过诱导ROS的产生导致胃癌细胞铁死亡,这是荜茇酰胺抗肿瘤的作用机制之一。Objective To study the effect and possible mechanism of piperlongumine(PIP)on iron death in gastric cancer cells HGC-27.Methods HGC-27 cells were interfered with PIP at a gradient concentration,and the cells were randomly divided into control group and PIP group.Control group was conventionally cultured cells,and the PIP groups were intervened with 5,10,and 20μmol·L^-1 PIP,respectively.Cell counting kit-8(CCK-8)was used to detect changes of cell viability,dichlorofluorescein(DCFH-DA)probe was used to detect reactive oxygen species(ROS)production,and Western blot was used to detect glutathione peroxide.The expression levels of glutathione peroxidase 4(GPX4),the cystine-glutamate reverse transporter subunit xCT protein,and metal ion transporter 1(DMT1)were tested.The kit was used to detect the intracellular malondialdehyde(MDA)content.HGC-27 cells were randomly divided into control group,PIP group,and NAC+PIP group by pretreatment with ROS inhibitor acetylcysteine(NAC).Control group was conventionally cultured cells.The NAC+PIP group was pretreated with 5μmol·L^-1 NAC for 12 h,and the PIP group and NAC+PIP group were intervened with 10μmol·L^-1.Cell viability,ROS production and related protein expression were also measured.Results After PIP intervention,the cell viability was significantly down-regulated,which was dose-dependent.After PIP intervention with a maximum of 20μmol·L^-1,the cell viability could be reduced to(50.34±0.13)%.Probe detection showed that intracellular ROS levels were increased,and intracellular MDA levels were also increased in a dose-dependent manner.After a maximum PIP intervention of 20μmol·L^-1,the MDA level could rise to(15.41±0.17)μmol·mL^-1.At the same time,the expressions of iron death key proteins GPX4 and xCT were down-regulated and DMT1 expression levels were up-regulated.NAC pretreatment can significantly inhibit iron death and increase cell viability.After 24 h,the cell viability of the NAC+PIP group was(65.42±0.41)%,and the cell viability of the PIP group was(58.21

关 键 词：荜茇酰胺 铁死亡 胃癌细胞 活性氧簇 

分 类 号：R28[医药卫生—中药学]