摘要
目的探讨吴茱萸碱(evodiamine,EVO)对人结肠癌细胞SW480迁移与侵袭的影响,并初步探究其潜在的分子机制。方法以CCK-8法测定不同浓度(0、0.375、0.75、1.5、3、6、12μmol/L)的EVO对SW480细胞处理不同时间(24、48、72 h)的增殖抑制作用;以划痕实验检测EVO对SW480细胞迁移的影响;Transwell实验检测EVO对SW480细胞侵袭的影响;Western blot检测IκBα、p-IκBα、NF-κB-p65、p-NF-κB-p65、MMP-9、MMP-2、MMP-13等蛋白的表达水平;免疫荧光实验检测EVO作用于SW480细胞后p-NF-κB-p65和MMP-9的蛋白表达情况;NF-κB-p65的激活剂LPS预处理SW480细胞后,Western blot检测EVO对IκBα、p-IκBα、NF-κB-p65、p-NF-κB-p65、MMP-9、MMP-2等蛋白表达的影响。结果 EVO呈时间及浓度依赖性抑制SW480细胞的增殖;经EVO作用后,SW480细胞的迁移能力下降(P<0.01),侵袭能力下降,呈浓度依赖性(P<0.01);EVO可以抑制NF-κB-p65信号通路(P<0.05);EVO对NF-κB-p65的抑制作用能被LPS削弱(P<0.05)。结论 EVO能抑制SW480细胞的迁移和侵袭,其机制可能是通过抑制IκBα的磷酸化,进而抑制NF-κB-p65的磷酸化,最终抑制下游基因MMP-9的表达。
Objective To investigate the effect of evodiamine(EVO)on the migration and invasion of colorectal cancer SW480 cells and explore the underlying mechanism.Methods SW480 cells were treated with different concentrations(0,0.375,0.75,1.5,3,6,12μmol/L)of EVO for 24,48,or 72 h,and the cell inhibition was detected using CCK-8 assay.SW480 cells treated with different concentrations of EVO for 24 h were examined for changes in migration and invasion using wound healing assay and Transwell assay and also for expression of IκBα,p-IκBα,NF-κB-p65,p-NF-κB-p65,MMP-9,MMP-2 and MMP-13 using Western blotting.Immunofluorescence staining was used to detect the expression of p-NF-κB-p65 and MMP-9 in SW480 cells treated with EVO for 24 h.The effects of lipopolysaccharide(LPS)challenge of SW480 cells prior to treatment with 1.5μmol/L EVO on the expressions of IκBα,p-IκBα,NF-κB-p65,p-NF-κB-p65,MMP-9 and MMP-2 were investigated using Western blotting.Results EVO inhibited the proliferation of SW480 cells in a time-and dose-dependent manner.EVO treatment also dose-dependently inhibited the migration ability(P<0.01)and invasion(P<0.01)of SW480 cells,and significantly inhibited NF-κB-p65 signaling(P<0.05).The inhibitory effect of EVO on NF-κB-p65 signaling was obviously attenuated by LPS challenge of the cells(P<0.05).Conclusion EVO can inhibit the expression of MMP-9 via suppressing NF-κB-p65 signaling,and then cause ultimately inhibition of the migration and invasion of SW480 cells.
作者
周鹏
吕艳伟
李静
陈地龙
ZHOU Peng;LYU Yanwei;LI Jing;CHEN Dilong(College of Public Health and Management,Chongqing Medical University,Chongqing,400016;Laboratory of Stem Cell and Tissue Engineering,Department of Histology and Embryology,College of Basic Medicine,Chongqing Medical University,Chongqing,400016;Chongqing Three Gorges Medical College,Chongqing,404120,China)
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2019年第6期549-555,共7页
Journal of Third Military Medical University
基金
国家自然科学基金面上项目(31271368)~~
关键词
吴茱萸碱
NF-κB-p65
MMP-9
结肠癌
迁移
侵袭
evodiamine
nuclear factor-κB-p65
matrix metalloproteinase-9
colorectal cancer
migration
invasion
