长链非编码RNA NR2F1-AS1靶向微小RNA-145-5p调控结肠癌细胞增殖、迁移和侵袭的分子机制

Molecular mechanism of lnc RNA NR2F1-AS1 targeting miR-145-5p to regulate colon cancer cell proliferation,migration and invasion

目的探讨长链非编码RNA(lncRNA)核受体亚族2F组成员1反义RNA(NR2F1-AS1)对结肠癌细胞增殖、迁移和侵袭的影响及作用机制。方法收集2015年3月至2017年5月青海省第五人民医院外科手术切除并经病理证实为原发性结肠腺癌组织25例,另留取距肿瘤边缘3 cm的癌旁组织(正常结肠组织)。实时荧光定量PCR(RT-qPCR)检测结肠癌组织和癌旁组织中NR2F1-AS1和微小RNA-145-5p(miR-145-5p)表达水平。以结肠癌细胞HCT116为研究对象,双荧光素酶报告基因实验验证NR2F1-AS1和miR-145-5p调控关系。转染NR2F1-AS1小干扰RNA或miR-145-5p模拟物至HCT116细胞,细胞计数试剂盒(CCK-8)和Transwell分别检测干扰NR2F1-AS1表达或过表达miR-145-5p对HCT116细胞增殖、迁移和侵袭的影响。结果与癌旁组织比较,结肠癌组织中NR2F1-AS1表达升高[(3.39±0.33)比(1.01±0.11),P<0.05],miR-145-5p表达降低[(0.55±0.05)比(1.00±0.08),P<0.05]。与共转染WT-NR2F1-AS1的miR-NC组比较,miR-145-5p组细胞荧光素酶活性降低P<0.05)。过表达NR2F1-AS1后细胞中miR-145-5p表达水平降低(P<0.05),干扰sNR2F1-AS1表达后细胞miR-145-5p表达水平升高(P<0.05)。干扰NR2F1-AS1表达或过表达miR-145-5p后,HCT116细胞光密度(OD)值、迁移和侵袭数降低(P<0.05)。干扰miR-145-5p表达逆转了干扰NR2F1-AS1表达对HCT116细胞OD值、迁移和侵袭数的影响。结论NR2F1-AS1在结肠癌组织中呈高表达,干扰NR2F1-AS1表达可能通过上调miR-145-5p表达来抑制结肠癌细胞恶性行为,是结肠癌治疗的潜在靶点。

Objective To investigate the effect and mechanism of lnc RNA NR2F1-AS1 on the proliferation,migration and invasion of colon cancer cells.Methods From March 2015 to May 2017,25 patients with primary colorectal adenocarcinoma confirmed by pathology and surgically resected in The Fifth People's Hospital of Qinghai Province were collected.Paracancental tissue(normal colon tissue)3 cm away from the tumor edge was also collected.The expression levels of NR2F1-AS1 and miR-145-5p in colon cancer tissues and adjacent tissues were detected by RT-qPCR.The colon cancer cell HCT116 was used as the research object,and the double luciferase reporter gene experiment verified the regulatory relationship between NR2F1-AS1 and miR-145-5p.NR2F1-AS1 small interfering RNA or miR-145-5p mimic was transfected into HCT116 cells.CCK-8 and Transwell were used to detect the effect of interfering with NR2F1-AS1 expression or over-expressing miR-145-5p on the proliferation,migration and invasion of HCT116 cells.Results Compared with adjacent tissues,the expression of NR2F1-AS1 in colon cancer tissues increased[(3.39±0.33)vs.(1.01±0.11),P<0.05],and the expression of miR-145-5p decreased[(0.55±0.05)vs.(1.00±0.08),P<0.05].Compared with the miR-NC group co-transfected with WT-NR2F1-AS1,the luciferase activity in miR-145-5p group was reduced(P<0.05).After over-expressing NR2F1-AS1,the expression level of miR-145-5p in cells decreased(P<0.05),and the expression level of miR-145-5p in cells increased after interfering with sNR2F1-AS1 expression(P<0.05).After interfering with sNR2F1-AS1 expression or over-expressing miR-145-5p,the OD value,HCT116 cells migration and invasion numbers were decreased(P<0.05).Interfering with miR-145-5p expression reversed the effect of interfering with NR2F1-AS1 expression on HCT116 cells OD value,migration and invasion numbers.Conclusion NR2F1-AS1 is highly expressed in colon cancer tissues.Interfering with NR2F1-AS1 expression may inhibit colon cancer cell proliferation,migration and invasion by up-regulating miR-145-5p expression,and is a potential target for colon cancer treatment.