Mechanisms of astrocyte elevated gene-1 promoting cancer metastasis through regulation of the micro-RNA-885-5p/ matrix metalloproteinase 9 signaling pathway in liver cancer

Abstract

Objective To investigate the relationship between astrocyte elevated gene-1 (AEG-1) and microRNA-885-5p, observe the influence of microRNA-885-5p on MHCC-97H migration and invasion, identify the target gene of microRNA-885-5p and uncover the mechanisms of AEG-1 promoting cancer metastasis. Methods The experimental study was adopted. The AEG-1 over-expressed plasmid constructed and the siRNA of silent AEG-1 synthesized were transiently transfected into MHCC-97H cells, respectively. For plasmid trans-fection, MHCC-97H cells tranfected with empty Vector NC were divided into group A as the control, and MHCC-97H cells tranfected with AEG-1 over-expressed plasmid were divided into group B. For AEG-1 siRNA transfection, MHCC-97H cells tranfected with negative siRNA were divided into group C as the control, and MHCC-97H cells tranfected with AEG-1 siRNA were divided into group D. The relative expressions of microRNA-885-5p in MHCC-97H cells of group A, B, C, D were measured by fluorescent quantitative polymerase chain reaction(PCR). For microRNA-885-5p simulator transfection, MHCC-97H cells tranfected with negative microRNA simulator were divided into group E as the control, and tranfected with microRNA-885-5p simulator were divided into group F. Transwell assay was used to evaluate MHCC-97H migration and invasion. Targetscan software was used to predict the target gene of microRNA-885-5p. MHCC-97H cells co-transfected with reporter plasmid of target gene and negative control siRNA were divided into group G as the control, and MHCC-97H cells co-transfected with reporter plasmid of target gene and microRNA-885-5p simulator were divided into group H. MHCC-97H cells co-transfected with reporter plasmid of target gene of binding region mutation and negative control siRNA were divided into group I as the control, MHCC-97H cells co-transfected with reporter plasmid of target gene of binding region mutation and microRNA-885-5p simulator were divided into group J. The luciferase activities of MHCC-97H cells in group G, H, I, J were tested. MHCC-97H cells transfected with negative control siRNA were divided into group K, and tranfected with microRNA-885-5p simulator were divided into group L. Western blot test was used to detect the relative expression of target gene of microRNA-885-5p. MHCC-97H cells transfected with siRNA of target gene of microRNA-885-5p were divided into group M, and transfected with negative control siRNA were divided into group N. Transwell assay was used to evaluate MHCC-97H migration and invasion in group M and group N. Measurement data with normal distribution were presented as ±s and comparison between groups was done using the t test. Results The relative expressions of microRNA-885-5p in MHCC-97H cells of group A and group B were (10.68±1.32)×10-4 and (5.02± 0.20)×10-4, respectively, showing significant difference between the 2 groups (t=7.357, P 0.05) . The relative expressions of MMP9 in MHCC-97H cells of group K and group L were 0.75±0.03 and 0.25±0.03, respectively, showing significant difference between the 2 groups (t=19.086, P<0.05). The results of MHCC-97H migration and invasion showed that the numbers of MHCC-97H cells in lower chambers of transwell plate were 1 210±163 and 1 192±170 in group M, 537±16 and 374±55 in group N, showing significant differences between the 2 groups (t=7.111, 7.916, P<0.05). Conclusions AEG-1 downregulates the expression of microRNA-885-5p, the target gene of which is MMP9. MicroRNA-885-5p inhabits the migration and invasion of hepatoma cells by negative regulation of MMP9. Key words: Liver neoplasms; Metastases; Astrocyte elevated gene-1; MicroRNA-885-5p; Matrix metalloproteinase 9