芒柄花黄素调控miR-182/FOXO3轴对子宫内膜癌细胞迁移及侵袭的影响  

Effect of formononetin on the migration and invasion of endometrial cancer cells by regulating the miR-182/FOXO3 axis

作　　者：梁殿迅[1] 郭哲[1] 朱军义[1] 许静[1] 付小玲[1] LIANG Dianxun;GUO Zhe;ZHU Junyi;XU Jing;FU Xiaoling(Henan Nanyang Central Hospital,Nanyang 473000,China)

机构地区：[1]河南省南阳市中心医院,473000

出　　处：《现代泌尿生殖肿瘤杂志》2023年第1期50-57,共8页Journal of Contemporary Urologic and Reproductive Oncology

摘　　要：目的探讨芒柄花黄素(FM)对子宫内膜癌细胞迁移及侵袭的影响,以及其作用机制。方法用0、12.5、25.0、50.0、100、200μmol/L FM处理人子宫内膜癌细胞株Ishikawa细胞48 h,流式细胞术检测细胞凋亡率;取生长状态良好的Ishikawa细胞,分别用25.0、50.0、100μmol/L FM处理,并命名为FM-L组、FM-M组、FM-H组,另取未处理的Ishikawa细胞作为对照组;利用细胞转染技术对处于对数生长期的Ishikawa细胞进行转染,分为miR-182 mimics组、miR-NC组、miR-182 inhibitor组、miR-NC inhibitor组、pcDNA-FOXO3组、pcDNA组,将miR-NC、miR-182 mimics转染Ishikawa细胞后再用100μmol/L FM处理,记为FM+miR-NC组、FM+miR-182 mimics组;Transwell测定细胞迁移及侵袭;Western Blot检测细胞中基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)、叉头转录因子O亚型3(FOXO3)蛋白表达;RT-PCR检测细胞中miR-182表达;双荧光素酶报告基因实验验证miR-182与FOXO3的靶向关系。结果与0μmol/L FM相比,12.5μmol/L、25.0μmol/L、50.0μmol/L、100μmol/L、200μmol/L FM均可促进Ishikawa细胞凋亡,且25.0μmol/L、50.0μmol/L、100μmol/L FM对Ishikawa细胞凋亡的促进作用最显著;与对照组比较,FM-L组、FM-M组和FM-H组的Ishikawa细胞迁移及侵袭数目、MMP-2及MMP-9蛋白和miR-182表达水平显著降低,FOXO3蛋白表达水平显著升高(P<0.05),且FM浓度越高,对应的变化趋势越明显;miR-182可靶向负调控FOXO3的表达;抑制miR-182表达或上调FOXO3表达均可抑制Ishikawa细胞迁移及侵袭数目和MMP-2、MMP-9蛋白表达(P<0.05);miR-182过表达逆转了FM(100μmol/L)对Ishikawa细胞迁移及侵袭的作用。结论FM可通过抑制miR-182并上调FOXO3的表达来抑制Ishikawa细胞的迁移和侵袭。Objective To investigate the effects of formononetin(FM)on the migration and invasion of endometrial cancer cells,and its mechanism of action.Methods Human endometrial carcinoma cell line Ishikawa cells were treated with 0,12.5,25.0,50.0,100,200μmol/L FM for 48 h,and apoptosis rate was detected by flow cytometry.Ishikawa cells in good growth condition were treated with 25.0,50.0,100μmol/L FM,named as FM-L group,FM-M group and FM-H group,and the untreated Ishikawa cells were taken as control group.The Ishikawa cells in logarithmic growth phase were transfected by cell transfection technology,and divided into miR-182 mimics group,miR-NC group,miR-182 inhibitor group,miR-NC inhibitor group,pcDNA-FOXO3 group,and pcDNA group.After transfecting miR-NC and miR-182 mimics into Ishikawa cells,they were treated with 100μmol/L FM,as FM+miR-NC group and FM+miR-182 mimics group.Transwell was used to measure cell migration and invasion.Western Blot was used to detect the expression of matrix metalloproteinase-2(MMP-2),matrix metalloproteinase-9(MMP-9)and forkhead transcription factor O subtype 3(FOXO3)proteins in cells.RT-PCR was used to detect the expression of miR-182 in cells.Dual luciferase reporter gene experiment was used to verify the targeting relationship between miR-182 and FOXO3.Results Compared with 0μmol/L FM,12.5μmol/L,25.0μmol/L,50.0μmol/L,100μmol/L,200μmol/L FM can promote the apoptosis of Ishikawa cells,and 25.0μmol/L,50.0μmol/L,100μmol/L FM had the most significant effect on the apoptosis of Ishikawa cells.Compared with the control group,the numbers of migration and invasion,and the expression levels of MMP-2,MMP-9 proteins and miR-182 in Ishikawa cells in the FM-L,FM-M,and FM-H groups were significantly reduced,and the expression level of FOXO3 protein was significantly increased(P<0.05),and the higher the FM concentration,the more obvious the corresponding trend.MiR-182 could target and negatively regulate the expression of FOXO3,and inhibiting miR-182 expression or up-regulating FOXO3 expressio

关 键 词：子宫内膜癌 芒柄花黄素 miR-182 叉头转录因子O亚型3 细胞迁移 细胞侵袭 

分 类 号：R285[医药卫生—中药学]