微小RNA-3915在肾癌中的表达及对肾癌细胞迁移和侵袭的影响

Expression of microRNA-3915 in renal cell carcinoma and its effect on migration and invasion of renal cancer cells

目的观察微小RNA-3915(miR-3915)在肾癌中的表达及其生物学意义。方法采用实时荧光定量聚合酶链式反应(qRT-PCR)检测14例肾癌及癌旁组织中miR-3915的表达量,同时检测正常肾小管上皮细胞株和肾癌细胞株中miR-3915的表达,以表达量最高的细胞株为实验对象。生物信息学预测软件预测并采用双荧光素酶报告基因验证miR-3915的靶基因。分别转染miR-3915抑制序列或阴性对照序列至肾癌细胞株,qRT-PCR检测miR-3915和靶基因mRNA的表达,Western blot检测细胞相应蛋白的表达。通过Transwell迁移、侵袭实验分别检测细胞的迁移和侵袭能力。结果相对于癌旁组织和正常肾小管上皮细胞,miR-3915在肾癌组织及肾癌细胞株中的表达明显上调(P<0.05),A498细胞表达量最高(P<0.01)。生物信息学预测并经双荧光素酶报告基因验证CMTM3是miR-3915的靶基因。miR-3915抑制序列可显著降低A498细胞miR-3915的表达(P<0.01),增加CMTM3mRNA的表达(P<0.01),使CMTM3、E-cadherin和β-catenin蛋白表达上调,Vimentin和Slug蛋白表达下调。A498细胞迁移和侵袭能力均降低(P<0.01)。结论 miR-3915在肾癌中的表达明显升高,通过抑制A498细胞中miR-3915的表达,可明显升高CMTM3基因的表达,抑制肾癌细胞的迁移和侵袭作用。

Objective To observe the expression of microRNA-3915 (miR-3915) in renal cell carcinoma and its biological significance. MethodsReal-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-3915 in 14 cases of renal cell carcinoma and adjacent tissues, and the expression of miR-3915 in normal renal tubular epithelial cells and renal cancer cell lines was detected. The cell line with the highest expression was used as the experimental object.Bioinformatics predicted and the dual luciferase reporter gene validated the target gene of miR-3915. The miR-3915 inhibitor sequence or miR-NC was transfected into renal cancer cell lines, qRT-PCR was used to detect the expression of miR-3915 and target gene mRNA, and the corresponding protein expression was detected by Western blot. Cell migration and invasion were tested by Transwell migration assay and Transwell invasion assay, respectively. ResultsCompared with paracancerous tissues and normal renal tubular epithelial cells, the expression of miR-3915 was significantly up-regulated in renal cell carcinoma and renal cancer cell lines ( P <0.05), and the expression in A498 cells was the highest ( P <0.01). Bioinformatics predicted and verified by the dual luciferase reporter gene that CMTM3 is a target gene of miR-3915. The inhibitor sequence of miR-3915 significantly decreased the expression of miR-3915 in A498 cells ( P <0.01), and the expression of CMTM3 mRNA was increased ( P <0.01). The expression of CMTM3, E-cadherin and β-catenin proteins were up-regulated, and the expression of Vimentin and Slug proteins were down-regulated. The migration and invasion ability of A498 cells were decreased ( P <0.01). ConclusionsThe expression of miR-3915 was significantly increased in renal cell carcinoma. By inhibiting the expression of miR-3915 in A498 cells, the expression of CMTM3 gene was significantly increased, then the migration and invasion of renal cancer cells were inhibited.