摘要
目的:研究土木香内酯(alantolactone)对食管癌EC9706细胞增殖、迁移、侵袭及凋亡的影响,并探讨可能的作用机制。方法:用不同浓度的土木香内酯处理食管癌EC9706细胞,以DMSO处理的EC9706细胞作为对照组。分别用结晶紫染色法、MTT法和克隆形成实验检测土木香内酯对EC9706增殖能力的影响;划痕愈合实验和Transwell小室法检测对EC9706细胞迁移和侵袭能力的影响;Hoechst 33258染色法检测对EC9706细胞凋亡的影响。蛋白印迹法检测侵袭转移相关蛋白基质金属蛋白酶7(matrix metalloproteinase-7,MMP-7)、MMP-9、波形蛋白(Vimentin)和Snail,以及凋亡相关蛋白Bax、Bcl-2、剪切型-caspase 3(cleaved-caspase 3)、caspase 3蛋白的表达水平。分别采用荧光素酶报告法和蛋白印迹法检测Wnt/β-catenin信号通路的活性及相关蛋白β-连环蛋白(β-catenin)、细胞周期相关蛋白D1(cyclin D1)和c-Myc的表达水平。结果:与对照组相比,土木香内酯可以明显抑制EC9706细胞的增殖、迁移和侵袭能力,并促进细胞凋亡(P值均<0.01),呈剂量和时间效应关系。不同浓度的土木香内酯均能上调凋亡相关蛋白Bax、cleaved-caspase 3和caspase 3的表达水平(P值均<0.01),下调抗凋亡蛋白Bcl-2、侵袭转移相关蛋白MMP-7、MMP-9、Vimentin和Snail的表达水平(P值均<0.01)。荧光素酶报告系统显示,土木香内酯能使T细胞因子/淋巴增强因子(T-cell factor/lymphoid enhancer factor,TCF/LEF)的转录活性明显下降(P<0.01);蛋白质印迹法检测结果显示土,木香内酯能明显抑制β-catenin、cyclin D1和c-Myc蛋白的表达水平(P值均<0.01)。结论:土木香内酯可能通过抑制Wnt/β-catenin信号通路的活性,抑制EC9706细胞的增殖、迁移和侵袭,并促进其凋亡。
Objective:To investigate the effects of alantolactone on proliferation,migration,invasion and apoptosis of esophageal cancer EC9706 cells,to explore the possible mechanisms.Methods:EC9706 cells were treated with different concentrations alantolactone,the proliferation was detected by crystal violet staining,MTT assay and colony forming assay,the migration and invasion abilities were detected by wound healing and Transwell assay,the apoptosis was detected by Hoechst 33258 staining assay.The expression levels of migration and invasion related proteins matrix metalloproteinase-7(MMP-7),MMP-9,Vimentin and Snail,apoptosis related proteins Bcl-2,Bax,caspase 3 and cleaved-caspase 3 were detected by Western blotting.The transcriptional activity of T-cell factor/lymphoid enhancer factor(TCF/LEF)was detected by Luciferase Reporter Gene system,then the expression levels of Wnt/β-catenin signaling pathway-related proteins c-Myc,cyclin D1 andβ-catenin were detected by Western blotting.Results:Compared with the control group,alantolactone could significantly inhibit the proliferation,migration and invasion of EC9706 cells,and could promote the apoptosis of EC9706 cells in a dose-time dependent manner(all P<0.01).Western blotting results showed that alantolactone could up-regulate the expression levels of Bax,cleaved-caspase 3 and caspase 3(all P<0.01),and down-regulate the expression levels of MMP-7,MMP-9,Bcl-2,Vimentin and Snail(all P<0.01).The transcription activity of TCF/LEF was significantly decreased(P<0.01),and it could inhibit the expression level ofβ-catenin,cyclin D1 and c-Myc(all P<0.01).Conclusion:The activity of Wnt/β-catenin signaling pathway may be inhibited by alantolactone,It can inhibit the proliferation,migration and invasion of EC9706 cells and promote its apoptosis.
作者
王紫洁
胡庆
陈欢
李婷
李强
邓明明
WANG Zijie;HU Qing;CHEN Huan;LI Ting;LI Qiang;DENG Mingming(Department of Gastroenterology,Affiliated Hospital of South west Med ical University,Luzhou 646000,Sichuan Province,China)
出处
《肿瘤》
CAS
CSCD
北大核心
2020年第10期665-675,共11页
Tumor
基金
四川省科技基金项目(编号:2016JY0101)
四川省泸州市科技基金项目(编号:2019-RCM-97)。
关键词
食管肿瘤
土木香内酯
细胞增殖
细胞凋亡
细胞运动
Esophageal neoplasms
Alantolactone
Cell proliferation
Apoptosis
Cell movement
