Effect of over-expressed miR-487A-3p on proliferation and invasion of renal cancer cells and its mechanism

Abstract

Objective To investigate whether microRNA-487a-3p (miR-487a-3p) can inhibit the proliferation and invasion of renal cancer cells by inhibiting the expression of its target gene TM4SF1 gene. Methods The miR-487a-3p mimics or miR-NC were transfected into renal cell carcinoma cells (ACHN) by LipofectamineTM 2000 and divided into experimental group and control group.The expression of miR-487a-3p after transfection was detected by real-time fluorescence quantitative polymerase chain reaction (qPCR). The cell cycle was evaluated by flow cytometry. MTT assay was used to detect cell proliferation. Transwell invasion assay was used to detect cell invasion. TM4SF1 wild-type (WT) and mutant 3'UTR-luciferase reporter vector were contructed. The dual luciferase reporter gene system was used to detect the effect of miR-487a-3p on the luciferase activity of TM4SF1 wild-type and mutant 3'UTR. The expression of TM4SF1 mRNA and protein in transfected cells were detected by qPCR and Western blot. Results The expression of miR-487a-3p in ACHN cells in the experimental and control groups was (33.12±3.35) and (1.04±0.18), respectively, with significant difference (t=9.553, P<0.001). The proportion of cells in experimental group(42.07±1.62) at G0-G1 phase was significantly higher than that(58.42±2.15) in control group (t=6.087, P=0.001). The proliferation activity of ACHN cell in the experimental group was significantly lower than the control group (P<0.05). The number of invasiveness of ACHN cells in the experimental group(114.70±20.38) was significantly lower than that(269.30±18.12) in the control group (t=5.670, P=0.001). Bioinformatics software and dual luciferase reporter assay showed that TM4SF1 was the target gene of miR-487a-3p (P<0.01). The expression of TM4SF1 mRNA in ACHN cells in experimental group(0.47±0.08) was significantly lower than that(1.02±0.10) in control group, and the difference was statistically significant (t=4.122, P=0.006). The expression of TM4SF1 protein in experimental ACHN cells was significantly lower than that in control group. Conclusions miR-487a-3p can inhibit the proliferation and invasion of ACHN cells in renal cell carcinoma by inhibiting the expression of its target gene TM4SF1, which is expected to become an important target for treatment of renal cell carcinoma in the future. Key words: Kidney Neoplasms; MicroRNAs; Cell Proliferation; Neoplasm Invasiveness