Abstract
BackgroundChemo-resistance of bladder cancer has been considered to be one of the serious issues to be solved. In this study, we revealed pivotal role of miR-424 in the regulation of CDDP sensitivity of bladder cancer cells.MethodsThe cytotoxicity of cisplatin and effect of miR-424 were assessed by flow cytometry and TUNEL. Transcriptional regulation of miR-424 by HIF-1α was assessed by Chromatin immunoprecipitation (ChIP). Effect of miR-424 on expression of UNC5B, SIRT4 (Sirtuin4) and apoptotic markers was measured by QRT-PCR and/or Western blot. The regulation of miR-424 for UNC5B and SIRT4 were tested by luciferase reporter assay. The 5637-inoculated nude mice xenograft model was used for the in vivo study. The clinical significance of miR-424 was demonstrated mainly through data mining and statistical analysis of TCGA.ResultsIn this study, we have found for the first time that cisplatin (CDDP) induces the expression of miR-424 in a HIF-1α-dependent manner under normoxia, and miR-424 plays a vital role in the regulation of CDDP resistance of bladder cancer cells in vitro. Mechanistically, we have found that UNC5B and SIRT4 are the direct downstream target genes of miR-424. CDDP-mediated suppression of xenograft bladder tumor growth was prohibited by the addition of miR-424, whereas ectopic expression of UNC5B or SIRT4 partially restored miR-424-dependent decrease in CDDP sensitivity of bladder cancer 5637 and T24 cells. Moreover, knockdown of UNC5B or SIRT4 prohibited CDDP-mediated proteolytic cleavage of PARP and also decreased CDDP sensitivity of these cells. Consistently, the higher expression levels of miR-424 were closely associated with the poor clinical outcome of the bladder cancer patients. There existed a clear inverse relationship between the expression levels of miR-424 and pro-apoptotic UNC5B or SIRT4 in bladder cancer tissues.ConclusionsCollectively, our current results strongly suggest that miR-424 tightly participates in the acquisition/maintenance of CDDP-resistant phenotype of bladder cancer cells through down-regulation of its targets UNC5B and SIRT4, and thus combination chemotherapy of CDDP plus HIF-1α/miR-424 inhibition might have a significant impact on hypoxic as well as normoxic bladder cancer cells.
Highlights
Chemo-resistance of bladder cancer has been considered to be one of the serious issues to be solved
Collectively, our current results strongly suggest that miR-424 tightly participates in the acquisition/ maintenance of CDDP-resistant phenotype of bladder cancer cells through down-regulation of its targets UNC5B and SIRT4, and combination chemotherapy of CDDP plus hypoxia-inducible factor-1α (HIF-1α)/miR-424 inhibition might have a significant impact on hypoxic as well as normoxic bladder cancer cells
The expression level of miR-424 is inversely correlated with those of UNC5B and SIRT4 in bladder cancer Since it has been described that miR-424 expression level is increased in certain human primary tumors [24,25,26], we Discussion In the current study, we have found for the first time that miR-424, a direct target of HIF-1α, decreases CDDP sensitivity of bladder cancer cells through down-regulation of pro-apoptotic UNC5B and SIRT4 (Fig. 6f)
Summary
Chemo-resistance of bladder cancer has been considered to be one of the serious issues to be solved. It has been indicated that both of the intrinsic and the acquired resistance to chemotherapy might cause the extremely poor clinical efficacy of anti-cancer drugs against bladder cancer [2, 3]. To overcome these serious issues, it is urgent to understand the precise molecular mechanisms how bladder cancer cells could acquire and/or maintain the chemo-resistant phenotype, which might contribute to the development of a novel therapeutic strategy against refractory bladder cancer [4]. It is likely that there could exist a promising HIF-1α-target gene(s) implicated in the regulation of chemo-sensitivity
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