The Study On The Growth Inhibitions Of Bladder Cancer By MiR-590-3p Target Regulating TFAM And Their Corresponding Mechanisms

Objectives:The miRNA acts as key factors in the regulation of gene expression,and the mitochondrial transcription factor A (TFAM) is required for mitochondrial DNA replication and transcription. However, the roles of miRNA and TFAM as well as their relationship in bladder cancer remain unclear. The aim of this study was to elucidate the roles of TFAM and miR-590-3p in bladder cancer cells and their corresponding molecular mechanisms.Methods:1The expression of miR-590-3p was detected by Real-time PCR, and the expression of TFAM was detected by Western-Blot in the bladder cancer, adjacent tissues and normal tissues.2The regulatory relation between miR-590-3p and TFAM was tested and verified by the Dual-Luciferase reporter assays.3The vectors of miR-590-3p and TFAM overexpression were constructed.4Effects of miR-590-3p and TFAM overexpression on proliferation, cell cycle, colony-formation efficiency and migration of5637cells were detected by MTT assays, flow cytometry assays and Trans-Well.5The expressions of PI3K, p-PI3k, AKT, P-AKT, MMP-2and MMP-9in cells which were transfected with miR-590-3p lentiviral vectors and TFAM over-expression vectors were detected, to seek the signal control paths of their regulatory relation.Results:1The bladder cancer tissues showed lower expression of miR-590-3p than normal tissue and adjacent tissues (P<0.05). Moreover, the higher stage of bladder cancer, the lower expression of miR-590-3p. In contrast the bladder cancer tissues showed higher expression of TFAM than normal tissue and adjacent tissues (P<0.05). Moreover, the higher stage of bladder cancer, the higher expression of TFAM.2The data showed that the renilla/firelfy value of lucifersae was significantly lower in miR-590-3p treatment cells after transfection with3’UTR of TFAM gene, while the renilla/firelfy value of lucifersae showed no difference after transfection with mutated3’UTR of TFAM compared with control. It suggested that miR-590-3p could down-regulated TFAM gene expression.3MTT data showed that miR-590-3p significantly markedly decreased the cell proliferation while TFAM could markedly promote cell proliferation. The cells transfected with miR-590-3p lentiviral vectors showed the highest percentage cells in G2stage, suggesting that the mitosis was blocked. For the cells transfected with TFAM over-expression vectors, we found that most of the cells were in G1and S stages and only a few cells were in G2stage, indicating that the cell division was active. The cells transfected with miR-590-3p lentiviral vectors showed the lowest colony-formation efficiency. For the cells transfected with TFAM over-expression vectors, we found the highest colony-formation efficiency. The miR-590-3p groups had the lowest transwell level among all groups meanwhile over-expression of TFAM significantly promoted cell migration.4The results indicated that the expressions of PI3K, p-PI3k, AKT, P-AKT, MMP-2and MMP-9was decreased in cells transfected with miR-590-3p lentiviral vectors, while increased in the cells transfected with TFAM over-expression vectors when compared with controls.Conclusions The negative control relationship between miR-590-3p and TFAM acts the anti-tumor role in bladder cancer, through lowering the PI3K/AKT path activity to down-regulate the expressions of MMP-2and MMP-9.