芹菜素对HeLa细胞miR-342-3p及其靶基因FOXM1表达和自我更新的影响  

Effects of Apigenin on Expressions of MiR-342-3p and its Target Gene FOXM1 and Self-Renewal in Cervical Cancer HeLa Cells

作　　者：赵霞[1] 乐伊婕 曹建国[2] 刘杰[1] 董巍檑[1] ZHAO Xia;LE Yijie;CAO Jianguo;LIU Jie;DONG Weilei(Department of Obstetrics and Gynecology,The First Affiliated Hospital,Hengyang Medical School,University of South China,Hengyang 421001;School of Pharmaceutical Sciences,Hunan Normal University,Changsha 410013)

机构地区：[1]南华大学第一附属医院妇产科,衡阳421001 [2]湖南师范大学药学院,长沙410013

出　　处：《湖南师范大学学报(医学版)》2024年第6期22-28,共7页Journal of Hunan Normal University(Medical Sciences)

基　　金：国家自然科学基金面上项目“蔓荆子总黄酮通过调控DNMT1和miR-342相互作用靶向E2F1抑制肝癌干性的机制研究”(82074075);湖南省卫健委科研项目“芹菜素激活miR-342-3p靶向FoxM1抑制宫颈癌干性研究”(202105012363)

摘　　要：目的:研究芹菜素(apigenin,API)是否通过上调miR-342-3p下调FOXM1表达水平,从而抑制宫颈癌HeLa细胞自我更新。方法:miR-342-3p抑制物、过表达FOXM1、亚细胞毒浓度API(1、3、10 μM)处理或API(10 μM)分别联合miR-342-3p抑制物或过表达FOXM1处理HeLa细胞。球形成法测定自我更新能力。实时定量PCR分析miRNAs和mRNA表达水平。蛋白免疫印迹分析蛋白表达水平。荧光素酶报告基因实验验证miR-342-3p是否直接靶向FOXM1。裸鼠异种移植瘤模型评估API对HeLa源性球形成细胞体内生长的作用。结果:亚细胞毒性的API(1、3、10 μM)以浓度依赖方式降低HeLa细胞球形成率[(32.70±2.48)% vs.(23.67±2.14)%、(16.50±1.66)%、(11.52±1.06)%,均P<0.05],下调CD49f和Sox2 蛋白表达水平,伴随提高miR-342-3p水平和降低FoxM1蛋白表达水平。此外,miR-342-3p抑制物或过表达FOXM1转染拮抗API的上述效应。口服API(50和100 mg/kg)抑制HeLa源性球形成细胞移植瘤生长[瘤重(0.66±0.11)g vs.(0.33±0.03)g、(0.17±0.02)g,均P<0.05]。荧光素酶报告基因实验证实miR-342-3p直接靶向FOXM1。结论:API经上调miR-342-3p下调靶基因FOXM1表达水平进而抑制宫颈癌HeLa细胞自我更新。Objective:To investigate whether apigenin(API)inhibits self-renewal by downregulating FOXM1 expression via miR-342-3p upregulation in cervical cancer HeLa cells.Methods HeLa cells transfected with miR-342-3p inhibitor or FOXM1 cDNA were treated with sub-cytotoxicity concentrations of API(1μM,3μM and 10μM)or in combination with API(10μM)treatment and miR-342-3p inhibitor or FOXM1 cDNA.Sphere formation assay was used to detect the self-renewal in HeLa cells.Quantitative real-time RT-PCR(qRT-PCR)was employed to detect miRNAs and mRNA expression levels.Immunoblotting was utilized to detect the expression levels of FoxM1,CD49f and Sox2 protein.The luciferase assay was applied to examine if FOXM1 is directly targeted by miR-342-3p.The efficacy of API on tumor growth in HeLa-derived sphere-forming cells(SFCs)in vivo was determined in a nude mouse xenograft model.Results The subcellular concentration of API(1μM;3μM and 10μM)decreased sphere formation rate[(32.70±2.48)%vs.(23.67±2.14)%,(16.50±1.66)%,(11.52±1.05)%,all of P<0.05)],down-regulated CD49f and Sox2 protein levels as well as elevated miR-342-3p levels and reduced FoxM1 protein expression levels in HeLa cells,in a dose-dependent tendency.Moreover,miR-342-3p inhibitor or FOXM1 cDNA could antagonize the aforementioned efficacy of API.In addition,oral API(50 and 100mg/kg)administration resulted in markedly reduced tumor growth[tumor weight:(0.66±0.11)g vs.(0.33±0.03)g,(0.17±0.02)g,all of P<0.05].Luciferase assay demonstrated FOXM1 is directly targeted by miR-342-3p.Conclusion:API reduces self-renewal capacity in cervical cancer HeLa cells by targeting FOXM1 via miR-342-3p upregulation.

关 键 词：HELA细胞 自我更新 芹菜素 miR-342-3p FOXM1 作用机制 

分 类 号：R737.3[医药卫生—肿瘤]