微RNA-340-5p靶向分化抑制因子3抑制人食管鳞状细胞癌细胞的增殖、迁移及侵袭

MicroRNA-340-5p inhibits proliferation,migration and invasion of human esophageal squamous cell carcinoma cells by targeting inhibitors of DNA binding/differentiation 3

目的探索miRNA-340-5p对分化抑制因子3(ID3)的调控机制及对食管鳞状细胞癌细胞生物学功能的影响。方法通过生物信息学分析、qRT-PCR及蛋白质印迹法初步验证人食管鳞状细胞癌Eca109细胞中miRNA-340-5p对ID3表达的影响,并检测12对食管鳞状细胞癌组织和癌旁正常组织中miRNA-340-5p和ID3的表达水平。采用双荧光素酶报告基因实验验证miRNA-340-5p对ID3的调控作用。采用CCK-8、平板克隆形成实验及Transwell实验考察miRNA-340-5p对Eca109细胞增殖、迁移和侵袭能力的影响。结果通过生物信息学分析及实验验证筛选出11个最有可能与ID3基因结合的miRNA,其中miRNA-340-5p在转录和翻译水平对ID3表达的下调作用最显著。双荧光素酶报告实验结果证实miRNA-340-5p能与ID3基因直接结合。相比癌旁正常组织,miRNA-340-5p在食管鳞状细胞癌组织中低表达、ID3 mRNA高表达(P均<0.01),且在癌组织中miRNA-340-5p与ID3 mRNA表达水平呈负相关(r=-0.71,P<0.01)。细胞功能学实验表明,miRNA-340-5p能够抑制Eca109细胞的增殖、迁移和侵袭(P均<0.01)。结论miRNA-340-5p靶向ID3基因抑制人食管鳞状细胞癌细胞的增殖、迁移及侵袭。

Objective To explore the mechanism of micro RNA(mi RNA)-340-5p in regulating inhibitors of DNA binding/differentiation 3(ID3)and its effect on the biological function of esophageal squamous cell carcinoma(ESCC)cells.Methods The effect of mi RNA-340-5p on ID3 expression in human ESCC Eca109 cells was preliminarily confirmed using bioinformatics analysis,quantitative real-time polymerase chain reaction and Western blotting.The expression levels of mi RNA-340-5p and ID3 were detected in 12 pairs of ESCC tissues and adjacent normal tissues.The regulatory effect of mi RNA-340-5p on ID3 was verified by dual-luciferase reporter assay.The effects of mi RNA-340-5p on the proliferation,migration and invasion of Eca109 cells were evaluated using cell counting kit 8,colony formation assay and Transwell assay.Results Eleven mi RNAs that most likely to bind to ID3 gene were identified through bioinformatics analysis and experimental verification,of which mi RNA-340-5p had the most significant effect on ID3 expression at the transcriptional and translation levels.Dual luciferase reporter assay confirmed that mi RNA-340-5p could directly bind to ID3 gene.The expression of mi RNA-340-5p was significantly lower and the expression of ID3 m RNA was significantly higher in the ESCC tissues compared with the adjacent normal tissues(both P<0.01),and the expression of mi RNA-340-5p was negatively correlated with the expression of ID3 m RNA in the ESCC tissues(r=-0.71,P<0.01).Cell function experiments showed that mi RNA-340-5p could inhibit the proliferation,migration and invasion of Eca109 cells(all P<0.01).Conclusion mi RNA-340-5p can inhibit the proliferation,migration and invasion of ESCC cells by targeting ID3 gene.