子宫内膜癌组织微小RNA-373表达情况及其对子宫内膜癌细胞增殖、迁移的影响研究

背景微小RNA-373 (miRNA-373)在多种恶性肿瘤(如肝癌、肺癌)的发生发展过程中具有重要作用,但其在子宫内膜癌(EC)发生发展中的作用尚不完全清楚。目的探讨EC组织miRNA-373表达情况及其对EC细胞增殖、迁移的影响。方法选取2012年6月-2013年6月在邯郸市妇幼保健院行手术治疗的64例EC患者的EC组织作为试验组,另选取同期在邯郸市妇幼保健院行子宫切除术的30例良性子宫肌瘤患者的正常组织作为对照组。采用实时荧光定量聚合酶链反应(qRT-PCR)检测两组miRNA-373表达情况,绘制Kaplan-Meier生存曲线以分析不同miRNA-373表达情况EC患者术后5年预后;分别比较转染miRNA-373模拟物(miRNA-373 mimic)及其阴性对照物(mimic-NC)、miRNA-373抑制剂(miRNA-373 inhibitor)及其阴性对照物(inhibitor-NC)的HEC-1B细胞大型肿瘤抑制基因2 (LATS2)表达情况、转染96 h后吸光度值、迁移细胞数量;采用双荧光素酶报告基因系统验证miRNA-373与LATS2间的靶标关系。结果 (1)试验组miRNA-373相对表达量高于对照组(P<0.05)。根据miRNA-373相对表达量平均值将试验组患者分为低表达组(n=26)和高表达组(n=38),Kaplan-Meier生存曲线显示,高表达组患者术后5年累积生存率为52.6%,低于低表达组的84.6%(P<0.05)。(2)转染miRNA-373 mimic的HEC-1B细胞LATS2相对表达量低于转染mimic-NC的HEC-1B细胞(P<0.05);转染miRNA-373 inhibitor的HEC-1B细胞LATS2相对表达量高于转染inhibitor-NC的HEC-1B细胞(P<0.05)。转染96 h后,转染miRNA-373 mimic的HEC-1B细胞吸光度值高于转染mimic-NC的HEC-1B细胞(P<0.05);转染miRNA-373 inhibitor的HEC-1B细胞吸光度值低于转染inhibitor-NC的HEC-1B细胞(P<0.05)。转染miRNA-373 mimic的HEC-1B细胞中迁移细胞数量多于转染mimic-NC的HEC-1B细胞(P<0.05);转染miRNA-373 inhibitor的HEC-1B细胞中迁移细胞数量少于转染inhibitor-NC的HEC-1B细胞(P<0.05)。(3)双荧光素酶报告基因系统检测结果显示,LATS2含有miRNA-373的潜在结合位点。野生型LATS2中miRNA-373相对荧光强度低于空载对照(P<0.05);突变型LATS2中miRNA-373相对荧光强度与空载对照比较,差异无统计学意义(P>0.05)。结论 EC组织miRNA-373呈高表达,其可通过直接靶向调节LATS2的表达而促进EC细胞增殖、迁移,进而影响EC患者预后。

Expression of miRNA-373 in Endometrial Carcinoma Tissue and Its Impact on Cell Proliferation and Migration

Background It is confirmed that microRNA-373(miRNA-373)plays an important role in the occurrence and development of kinds of tumors(such as liver cancer and lung cancer),however its role in the occurrence and development of endometrial cancer(EC)is not yet clear so far. Objective To analyze the expression of miRNA-373 in EC tissue and its impact on cell proliferation and migration. Methods EC tissues obtained from 64 patients underwent surgical resection were selected as experiment group in Handan Maternal and Child Health Hospital from June 2012 to June 2013,meanwhile normal tissues obtained from 30 benign uterine fibroids patients underwent hysterectomy were selected as control group. Real-time fluorescence quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression of miRNA-373 in the two groups,and Kaplan-Meier survival curve was drawn to analyze the prognosis in EC patients with different expression of mi RNA-373;comparison of expression of LATS2,optical density 96 hours after transfection and number of migratory cells was conducted between mi RNA-373 mimic and mimic-NC(negative control)transfected HEC-1 B cells,between mi RNA-373 inhibitor and inhibitor-NC(negative control)transfected HEC-1 B cells,respectively,moreover dual luciferase reporter gene system was used to verify the target relationship between mi RNA-373 and LATS2. Results (1)Relative expression quantity of mi RNA-373 in experiment group was statistically significantly higher than that in control group(P<0.05). Of the 64 patients with EC,26 cases were with low expression of mi RNA-373,the other 38 cases were with high expression of mi RNA-373;Kaplan-Meier survival curve showed that,the postoperative 5-year cumulative survival rate was 52.6% in EC patients with high expression of mi RNA-373,which was statistically significantly lower than that in EC patients with low expression of mi RNA-373 of 84.6%(P<0.05).(2)Relative expression quantity of LATS2 in HEC-1 B cells transfected with mi RNA-373 mimic was statistically significantly lower than that in HEC-1 B cells transfected with mimic-NC(P<0.05),while relative expression quantity of LATS2 in HEC-1 B cells transfected with mi RNA-373 inhibitor was statistically significantly higher than that in HEC-1 B cells transfected with inhibitor-NC(P<0.05). Optical density in HEC-1 B cells transfected with mi RNA-373 mimic was statistically significantly higher than that in HEC-1 B cells transfected with mimic-NC 96 hours after transfection,while optical density in HEC-1 B cells transfected with mi RNA-373 inhibitor was statistically significantly lower than that in HEC-1 B cells transfected with inhibitor-NC(P<0.05). Number of migratory cells in HEC-1 B cells transfected with mi RNA-373 mimic was statistically significantly more than that in HEC-1 B cells transfected with mimic-NC(P<0.05),while number of migratory cells in HEC-1 B cells transfected with mi RNA-373 inhibitor was statistically significantly less than that in HEC-1 B cells transfected with inhibitor-NC(P<0.05).(3)Dual luciferase reporter gene system detection results showed that,there was potential binding site of mi RNA-373 in LATS2. Relative intensity of fluorescence of mi RNA-373 in wild type of LATS2 was statistically significantly lower than that in blank-control(P<0.05),however there was no statistically significant differencein relative intensity of fluorescence of mi RNA-373 between mutant type of LATS2 and blank-control(P>0.05). Conclusion Expression of mi RNA-373 is high in EC tissue,and it may promote the cell proliferation and migration though direct targeting regulation of LATS2,and then affect the prognosis in patients with EC.