【摘要】 目的分析miR-144-3p对肺癌细胞迁移和侵袭的影响,并探讨其可能机制。方法采用实时荧光定量PCR(QPCR)检测肺癌NCI-H460细胞、正常支气管上皮细胞HBE中miR-144-3p、ONECUT2的表达;采用脂质体法将miR-144-3p mimics(miR-144-3p组)、miR-NC(miR-NC组)、si-con(si-con组)、si-ONECUT2(si-ONECUT2组)、miR-144-3p mimics和pcDNA(miR-144-3p+pcDNA组)、miR-144-3p mimics和pcDNA-ONECUT2(miR-144-3p+pcDNA-ONECUT2组)转染至NCI-H460细胞;Transwell法检测各组细胞的迁移、侵袭能力;Western blotting检测各组ONECUT2、E-cadherin、N-cadherin、Vimentin蛋白表达;双荧光素酶报告实验检测miR-144-3p和ONECUT2的靶向关系。结果与HBE组比较,NCI-H460组细胞中miR-144-3p表达显著降低,ONECUT2表达显著升高(P<0.05)。过表达miR-144-3p、沉默ONECUT2均可明显抑制NCI-H460细胞的迁移和侵袭能力,上调E-cadherin蛋白表达,下调N-cadherin、Vimentin蛋白表达;miR-144-3p可抑制野生型ONECUT2 3’-UTR报告基因载体的荧光素酶活性,而对突变型ONECUT2 3’-UTR-MUT的荧光素酶活性无影响。过表达ONECUT2可逆转miR-144-3p对NCI-H460细胞的EMT相关蛋白表达。结论 miR-144-3p过表达可抑制肺癌细胞的迁移和侵袭能力,其机制可能与靶向ONECUT2抑制EMT有关,可为肺癌的治疗提供参考。更多还原

【Abstract】 Objective To analyze the effect of miR-144-3 p on the migration and invasion of lung cancer cells and explore its possible mechanism.Real-time fluorescence quantitative polymerase chain reaction(QPCR) was used to detect the expression of miR-144-3 p and ONECUT2 in lung cancer NCI-H460 cells and normal bronchial epithelial cells HBE.miR-144-3 p mimics(miR-144-3 p group), miR-NC(miR-NC group), si-con(si-con group), si-ONECUT2(si-ONECUT2 group), miR-144-3 p mimics and pcDNA(miR-144-3 p+pcDNA group), miR-144-3 p mimics and pcDNA-ONECUT2(miR-144-3 p+pcDNA-ONECUT2 group) were transfected into NCI-H460 cells by liposome method.Transwell assay was used to detect the migration and invasion ability of cells in each group. The protein expression of ONECUT2, E-cadherin, N-cadherin, and Vimentin of each group were detected by Western blotting.Dual luciferase reporter gene assay was used to detect the targeting relationship between miR-144-3 p and ONECUT2. Results Compared with HBE group, the expression of miR-144-3 p and ONECUT2 in NCI-H460 group were significantly decreased and increased(P<0.05).Overexpression of miR-144-3 p or silence of ONECUT2 could significantly inhibit the migration and invasion of NCI-H460 cells, up-regulate E-cadherin protein expression, down-regulate N-cadherin and Vimentin protein expression.MiR-144-3 p inhibited the luciferase activity of wild-type ONECUT2 3’-UTR reporter gene vector, but had no effect on the luciferase activity of mutant ONECUT2 3’-UTR-MUT.Overexpression of ONECUT2 reversed the expression of EMT-related protein in NCI-H460 cells by miR-144-3 p. Conclusion Overexpression of miR-144-3 p can inhibit the migration and invasion of lung cancer cells. The mechanism may be related to the inhibition of EMT by targeting ONECUT2, which can provide a reference for the treatment of lung cancer.更多还原