超微氧化铁纳米粒子通过强化自噬机制抑制人肝细胞癌HepG2细胞的迁移和侵袭  

Ultrasmall iron oxide nanoparticles inhibit the migration and invasion of human hepatocellular carcinoma HepG2 cells by enhancing autophagy

作　　者：陈汉仁 蒋淑莲[2] 张鹏 任中玉 陈剑娇 文剑 CHEN Hanren;JIANG Shulian;ZHANG Peng;REN Zhongyu;CHEN Jianjiao;WEN Jian(School of Pharmacy,Guilin Medical University,Guilin 541040,Guangxi,China;Department of Integrated Chinese and Western Medicine,the Second Hospital of Nanjing,Nanjing 210003,Jiangsu,China;School of Clinical Medicine,Guilin Medical University,Guilin 541040,Guangxi,China;Department of Neurology,Affiliated Hospital of Guilin Medical College,Guilin 541001,Guangxi,China)

机构地区：[1]桂林医学院药学院,广西桂林541040 [2]南京市第二医院中西医结合科,江苏南京210003 [3]桂林医学院临床医学院,广西桂林541040 [4]桂林医学院附属医院神经内科,广西桂林541001

出　　处：《中国肿瘤生物治疗杂志》2021年第8期783-789,共7页Chinese Journal of Cancer Biotherapy

基　　金：国家自然科学基金地区基金资助项目(No.32060228);广西自然科学基金资助项目(No.2017GXNSFAA198112,No.2019GXNSFAA245077);广西研究生教育创新计划资助项目(No.YCSW2019214,No.YCSW2020225);桂林市科学研究与技术开发计划资助项目(No.20190219-2)。

摘　　要：目的:探讨超微氧化铁纳米粒子(ultrasmall iron oxide nanoparticle,USIONP)对人肝细胞癌HepG2细胞迁移和侵袭的影响及其可能的机制。方法:采用粒径分析仪和透射电镜分别分析USIONP的水合粒径和核心粒径,Zeta电位和胶体稳定性实验分析USIONP的分散性及其稳定性以鉴定USIONP的成功制备;用不同质量浓度USIONP(0、50、100、200μg/ml)或200μg/ml USIONP+5 mmol/L 3-MA(自噬抑制剂)联合处理HepG2细胞,CCK-8法检测HepG2细胞的增殖活力,Transwell法检测细胞的迁移和侵袭能力,WB实验检测自噬标志物Beclin1、LC3、p62的表达,2’,7’-二氯二氢荧光素二醋酸(DCFH-DA)法测定细胞内活性氧(ROS)水平,铁离子比色法检测细胞内铁离子水平。结果:USIONP的平均水合粒径为(37.86±12.90)nm、核心粒径约10 nm,Zeta电位为–23.8 mV,有良好的水溶分散性,证实了USIONP的成功制备。随USIONP质量浓度升高和处理时间延长,HepG2细胞的增殖活力明显降低(均P<0.05);与对照组相比,200μg/ml USIONP处理HepG2细胞24 h后,迁移、侵袭细胞数量均显著减少(均P<0.05),而3-MA能够部分抵消上述影响(均P<0.05)。与对照组相比,100、200μg/ml USIONP处理组的HepG2细胞中Beclin1和LC3Ⅱ蛋白相对表达水平均显著升高(均P<0.05),而p62蛋白表达水平下降(均P<0.05);200μg/ml USIONP可显著提高细胞内ROS水平与铁离子水平,而加入3-MA可阻断其作用(均P<0.05)。结论:USIONP能促进HepG2细胞发生自噬,而自噬通路激活后降解USIONP释放铁离子和导致细胞ROS水平升高,从而抑制HepG2细胞的迁移和侵袭。Objective:To explore the effects of ultrasmall iron oxide nanoparticles(USIONPs)on the migration and invasion of human hepatocellular carcinoma HepG2 cells and its possible mechanism.Methods:The hydrate particle size and core particle size of USIONPs were analyzed by particle analysis device and transmission electron microscope,respectively.The dispersity and stability of USIONPs were characterized by Zeta potential and colloid stability analysis,respectively,to identify the successful prepaation of USIONPs.Different concentrations of USIONPs(0,50,100,200μg/ml)or 200μg/ml USIONPs+5 mmol/L 3-MA(autophagy inhibitor)were used to treat human liver cancer HepG2 cells;then,CCK-8 assay was used to detect the proliferation viability of HepG2 cells;Transwell method was used to detect the migration and invasion ability of HepG2 cell;WB experiment was applied to detect the expression of autophagy markers Beclin1,LC3 and p62;2',7'-dichlorodihydrofluorescein diacetic acid(DCFH-DA)method was used to determine the intracellular reactive oxygen species(ROS)level,and the iron ion colorimetric method was used to detect the iron ion level in the cells.Results:The average hydrate particle size of USIONPs was(37.86±12.90)nm,the core particle size was about 10 nm,and the Zeta potential was–23.8 mV,and confirmed the successful preparation of USIONP.The USIONPs had good water solubility and dispersibility.As the mass concentration of USIONPs increased and the incubation time prolonged,the proliferation viability of HepG2 cells showed a decreasing trend.Compared with the control group,after incubating HepG2 cells with 200μg/ml USIONPs for 24 h,the numbers of migrated and invaded cells were significantly reduced(all P<0.05),and the addition of 3-MA could partially offset the above effects(P<0.05).Compared with the control group,the relative protein expression levels of beclin1 and LC3Ⅱin HepG2 cells in the 100 and 200μg/ml USIONPs treatment groups increased significantly(all P<0.05),while the p62 protein expression decreased sign

关 键 词：氧化铁纳米粒子 肝细胞癌 HEPG2细胞 迁移 侵袭 自噬 

分 类 号：R735.7[医药卫生—肿瘤] R730.5[医药卫生—临床医学]