长链非编码RNA LINC00641通过miR-181d-5p/FOXP1调控结肠癌细胞增殖

Proliferation regulation on lncRNA LINC00641 through miR-181d-5p/FOXP1 in colon cancer cell

目的分析长链非编码RNA(LncRNA)LINC00641在结肠癌细胞HCT-116中的表达特征,探讨LINC00641通过微小RNA-181d-5p(miR-181d-5p)/叉头框蛋白P1(FOXP1)对细胞增殖的调控作用。方法选择人结肠癌细胞系HCT-116、人正常结肠上皮细胞系CCD841,应用实时荧光定量PCR检测LINC00641和miR-181d-5p的表达。将未进行转染的HCT-116细胞设为空白对照组,并构建转染pc-DNA3.1的空载体转染组、转染pc-DNA3.1-LINC00641的pc-DNA-LINC00641组、转染pc-DNA3.1-LINC00641-siRNA的si-LINC00641组,转染miR-181d-5p inhibitor的miR-181d-5p inhibitor组、转染miR-181d-5p mimic的miR-181d-5p mimic组、转染siRNA-FOXP1的si-FOXP1组。应用CCK-8实验检测细胞增殖活性,应用Western blot检测增殖细胞核抗原(PCNA)的表达,应用双荧光素酶报告基因实验观察LINC00641和miR-181d-5p、miR-181d-5p和FOXP1的靶向关系。结果结肠癌细胞系中LINC00641的表达量明显低于正常结肠上皮细胞系,miR-181d-5p的表达量明显高于正常结肠上皮细胞系。与空白对照组和空载体转染组比较,pc-DNA-LINC00641组的细胞增殖活性明显降低,si-LINC00641组细胞增殖活性明显增高。双荧光素酶报告基因实验显示LINC00641和miR-181d-5p、miR-181d-5p和FOXP1具有靶向关系。挽救实验显示沉默FOXP1可以部分逆转过表达LINC00641和沉默miR-181d-5p对细胞增殖的抑制作用及PCNA的低表达状态。结论结肠癌细胞系中LINC00641的表达下调,LINC00641通过miR-181d-5p/FOXP1途径抑制结肠癌细胞的增殖。

Objective To analyze the expression characteristics of long non coding RNA(lncRNA)LINC00641 in colon cancer cell line HCT-116,and to explore the regulation of LINC00641 on cell proliferation through microRNA-181d-5p(miR-181d-5p)/forkhead box protein 1(FOXP1).Methods Human colon cancer cell line HCT-116 and human normal colon epithelial cell line CCD841 were selected.Expressions of LINC00641 and miR-181d-5p were detected by real time fluorescent quantitative PCR in two cell lines.We constructed blank control group(without transfection),empty vector transfection group(pc-DNA3.1),pc-DNA-LINC00641 group(pc-DNA3.1-LINC00641 transfection),si-LINC00641 group(pc-DNA3.1-LINC00641-siRNA transfection),miR-181d-5p inhibitor group,miR-181d-5p mimic group,and si-FOXP1 group(siRNA-FOXP1 transfection).Cell proliferation activity was detected by CCK-8 assay.PCNA was detected by Western blot method.Targeting relationship between LINC00641 and miR-181d-5p,miR-181d-5p and FOXP1 were analyzed by dual luciferase reporter gene assay.Results Expression of LINC00641 was significantly lower in colon cancer cell line than that in normal colon epithelial cell line,the expression of miR-181d-5p was significantly higher than that in normal colon epithelial cell line.Compared with blank control group and empty vector transfection group,the proliferation activity of pc-DNA-LINC00641 group was decreased,while that of si-LINC00641 group was increased.Targeted relationship of LINC00641 and miR-181d-5p,miR-181d-5p and FOXP1 were confirmed by dual luciferase reporter gene experiment.The rescue experiment showed that silencing FOXP1 could partially reverse the inhibition of cell proliferation by overexpression of LINC00641 and miR-181d-5p and the low expression of PCNA.Conclusion Lower expression of LINC00641 is found in colon cancer cell line.LINC00641 inhibits the proliferation through miR-181d-5p/FOXP1 pathway in colon cancer cells.