微小核糖核酸-146a通过靶向调控B-细胞淋巴瘤因子11A对子宫内膜样腺癌细胞增殖、迁移和上皮间质转化的影响

The effect of miR-146a on the proliferation,migration and EMT of endometrioid adenocarcinoma cells through targeted regulation of BCL11A

目的探讨微小核糖核酸-146a(miR-146a)对子宫内膜样腺癌(EA)细胞增殖、迁移和上皮间质转化(EMT)的影响。方法选取2019年4月至2021年4月于河北北方学院附属第一医院接受手术治疗的45例子宫内膜癌(EC)患者的癌组织及邻近正常组织作为研究对象,根据免疫组化染色结果分为EA组(n=31)和非EA组(n=14);体外培养人子宫内膜上皮HEECs细胞和EA细胞株Ishikawa、HEC-1A,并分别转染mimics NC、miR-146a mimics进行分组;另购得20只雄性BALB/c裸鼠进行动物实验。采用实时荧光定量聚合酶链反应(RT-qPCR)测定各组织和细胞中miR-146a的表达水平;分别采用四甲基偶氮唑蓝(MTT)比色实验、集落形成实验、Transwell实验测定mimics NC组与miR-146a mimics组细胞的活力、增殖、迁移、侵袭能力;采用Western blot分析mimics NC组和miR-146a mimics组细胞中EMT相关蛋白、B-细胞淋巴瘤因子11A(BCL11A)的表达情况;采用膜联蛋白V-异硫氰酸荧光素和碘化丙啶(Annexin V-FITC/PI)双染法测定细胞的凋亡情况;采用荧光素酶报告基因检测验证miR-146a对BCL11A的靶向调控作用;采用裸鼠异种移植验证miR-146a对体内EA发生的影响。结果EC组织和细胞中的miR-146a水平明显降低(P<0.05)。与mimics NC组相比,miR-146a mimics组Ishikawa、HEC-1A细胞的miR-146a、E-cadherin、α-catenin水平及凋亡率明显升高,N-cadherin、Vimentin、BCL11A水平及细胞活力、克隆数、迁移数目、侵袭数目、荧光素酶活性明显下降(P<0.05)。动物实验结果显示,miR-146a mimics组的肿瘤体积明显小于mimics NC组(P<0.05)。结论过表达miR-146a可抑制EA细胞的增殖、迁移和EMT,促进细胞凋亡,其机制可能是通过靶向下调BCL11A实现的。

Objective To explore the effects of microribonucleic acid-146 a(miR-146 a)on the proliferation,migration and epithelial-mesenchymal transition(EMT)of endometrioid adenocarcinoma(EA)cells.Methods 45 cases of endometrial cancer(EC)patients who underwent surgical treatment in the First Affiliated Hospital of Hebei North University from April 2019 to April 2021 were selected as the research objects.According to the immunohistochemical staining results,they were divided into EA group(n=31)and non-EA group(n=14).Human endometrial epithelial HEECs cells and EA cell lines Ishikawa and HEC-1 A were cultured in vitro and transfected with mimics NC and miR-146 a mimics for grouping.In addition,20 male BALB/c nude mice were purchased for animal experiments.The expression level of miR-146 a in tissues and cells was determined by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).The activity,proliferation,migration and invasion of mimics NC group and miR-146 a mimics group were determined by MTT assay,colony formation assay and Transwell assay,respectively.Western blot was used to analyze the expression of EMT-related protein and B-cell lymphoma factor 11 A(BCL11 A)in the mimics NC group and miR-146 a mimics group.Apoptosis of the cells was determined by Annexin V-FITC/PI double staining.Luciferase reporter gene assay was used to verify the targeted regulation of miR-146 a on BCL11 A.The effect of miR-146 a on EA in vivo was verified by xenotransplantation in nude mice.Results The level of miR-146 a in EC tissues and cells was significantly reduced(P<0.05).Compared with the mimics NC group,the miR-146 a mimics group Ishikawa and HEC-1 A cells have significantly increased levels of miR-146 a,E-cadherin,α-catenin,and apoptosis rate,and have significantly decreased levels of N-cadherin,Vimentin,BCL11 A,cell viability,number of cell clones,number of cell migration,number of invaded cells,and luciferase activity(P<0.05).Animal experiments showed that the tumor volume of the miR-146 a mimics group was significantly lower than that of the mimics NC group(P<0.05).Conclusions Overexpression of miR-146 a can inhibit the proliferation,migration and EMT of EA,and promote cell apoptosis.The mechanism may be achieved by targeted down-regulation of BCL11 A.