摘要
目的:探讨miR-27a对皮肤鳞状细胞癌(CSCC)细胞增殖、侵袭、迁移的影响及其与叉头框Gl(FOXG1)的靶向关系.方法:收集 30 例 CSCC 组织及 30 例正常皮肤组织.培养 A431 细胞,将 miR-NC、miR-27a mimics、si-NC、si-miR-27a、Scramble、si-FOXG1、Vector、OE-FOXG1 质粒分别转染至细胞,记为 miR-NC 组、miR-27a 组、si-NC 组、si-miR-27a 组、Scramble 组、si-FOXG1 组、Vector 组及 FOXG1 组;将 si-FOXG1、Scramble 质粒分别转染至 miR-27a 组细胞,记为 miR-27a+Scramble 组、miR-27a+si-FOXG1组.四甲基偶氮唑蓝(MTT)检测细胞增殖能力,Transwell实验检测细胞侵袭、迁移能力,荧光定量聚合酶链反应(RT-PCR)检测细胞、组织中FOXG1基因mRNA及miR-27a表达水平,Western blot检测细胞或组织FOXG1蛋白表达水平,TargetScan在线网站预测miR-27a与FOXG1结合位点,双荧光素酶报告基因实验验证miR-27a与FOXG1的靶向关系.结果:CSCC组织FOXG1基因mRNA、miR-27a表达水平高于正常皮肤组织(P＜0.05),CSCC组织FOXG1基因mRNA、miR-27a表达水平呈正相关(r=0.801,P=0.000).上调miR-27a、FOXG1可促进细胞的增殖、侵袭、迁移,沉默miR-27a、FOXG1可抑制细胞的增殖、侵袭、迁移及EMT.下调FOXG1逆转了过表达miR-27a对细胞的增殖、侵袭、迁移的促进作用.miR-27a可正性调控FOXG1表达.结论:miR-27a、FOXG1在CSCC组织中高表达,miR-27a正性调控FOXG1促进CSCC细胞的增殖、侵袭、迁移.
Abstract
Objective:To investigate the effects of miR-27a on Proliferation,invasion and migration of cutaneous squamous cell carcinoma(CSCC)cells and its targeting relationship with forkhead box G1(FOXG1).Methods:30 cases CSCC tissues and 30 cases normal skin tissues were collected.A431 cells were cultured.miR-NC,miR-27a mimics,si-NC,si-miR-27a,Scramble,si-FOXG1,Vec-tor,and OE-FOXG1 plasmids were transfected into the cells and defined as miR-NC group,miR-27a group,si-NC group,si-miR-27a group,Scramble group,si-FOXG1 group,Vector group and FOXG1 group.The si-FOXG1 and Scramble plasmids were transfected into the cells of miR-27a group and recorded as miR-27a+Scramble group and miR-27a+si-FOXG1 group.The cell Proliferation ability was detected by tetramethylazole blue(MTT).The cell invasion and migration ability were detected by Transwell assays.FOXG1 gene mRNA and miR-27a level were detected by fluorescence quantitative polymerase chain reaction(RT-PCR).FOXG1 protein level were detected by Western blot.TargetScan online website was used to predict the binding site of miR-27a and FOXG1.Dual luciferase reporter gene assay was used to verify the targeting relationship between miR-27a and FOXG1.Results:The levels of FOXG1 gene mRNA and miR-27a in CSCC tissues were higher than normal skin tissues(P＜0.05),and the levels of FOXG 1 gene mRNA in CSCC tissues were positively correlated with and miR-27a level(r=0.801,P=0.000).Up-regulation of miR-27a and FOXG1 promoted cell prolifera-tion,invasion and migration,and silencing of miR-27a and FOXG1 inhibited cell proliferation,invasion,migration and EMT.Down-reg-ulation of miR-27a reversed the the promotive effect of cell Proliferation,invasion and migration by overexpression of FOXG 1.miR-27a positively regulated FOXG1 expression.Conclusion:miR-27a and FOXG1 are highly expressed in CSCC tissues.miR-27a positively regulates FOXG1 to promote proliferation,invasion,and migration of CSCC cells.
基金项目
新疆维吾尔自治区科技支疆计划项目(2019E0289)
NETL
NSTL
万方数据