Suppression of HAX-1 induced by miR-325 resensitizes bladder cancer cells to cisplatin-induced apoptosis

摘要

OBJECTIVE: MicroRNA-325 (miR-325) is a tumor suppressor in some cancers. However, the role of miR-325 in determining the chemosensitivity to cancer cells is still not clear. The aim of this study was to investigate the effect of miR-325 on reversing the cisplatin resistance of bladder cancer. MATERIALS AND METHODS: Cisplatin-resistant 5637 and T24 bladder cancer cell lines (5637/R and T24/R) were established through long term exposure of them to cisplatin. 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assays were performed to evaluate the viability of 5637, 5637/R, T24, and T24/R cells. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) was used to examine the expression of miR-325 in these cell lines. The regulatory mechanism was confirmed by Western blot analysis and Luciferase reporter assays. After treatment with miR-325 and cisplatin, mitochondrial membrane potential (MMP) and apoptosis were measured using flow cytometry. Expression of hematopoietic cell-specific protein 1-associated protein X-1 (HAX-1) and activation of caspase-9, caspase-7, and caspase-3 were detected by Western blotting. RESULTS: We found the downregulation of miR-325 in 5637/R and T24/R cells compared to their parental 5637 and T24 cells. Moreover, overexpression of miR-325 in cisplatin-resistant bladder cancer cells was found to increase the cytotoxicity of cisplatin to them. However, transfection with HAX-1 plasmid can abolish the effect of miR-325 on cisplatin. We, then, showed that overexpression of miR-325 suppressed the expression of HAX-1. Thus, miR-325 promoted the mitochondria collapse and cisplatin-induced apoptosis in bladder cancer cells. CONCLUSIONS: Downregulation of miR-325 is responsible for the development of cisplatin resistance in bladder cancer. Overexpression of miR-325 may represent a novel strategy to reverse the chemoresistance of bladder cancer.