MicroRNA-218 Inhibits Bladder Cancer Cell Proliferation,Migration And Invasion By Targeting BMI-1

Background:Bladder cancer is one of the common urological malignancies.miRNAs are recognized as a class of small noncoding RNAs and have emerged as important gene regulators in tumorigenesis.Growing evidence suggested that miR-218 was a tumor suppressor in many human cancers.However,its underlying role in bladder cance remains unclear.The aim of this study was to explore the effect of miR-2J8 on the proliferation,migration and invasion of BCa cells.Methods:The expression of miR-218 in 27 pairs of BCa tissues and normal adjacent tissues were quantified by qRT-PCR.The lentivirus of pHBLV-U6-ZsGreen-Puro-miR-218 or pHBLV-U6-ZsGreen-Puro-vector was constructed.Both lentivirus were transfected into bladder cancer T24 and EJ cell lines respectively.The expression of miR-218 and BMI-1 in the transfected T24 and EJ cell lines were detected by qRT-PCR.The protein levels of BM1-1,PTEN,AKT and pAKT in cell lines were detected by western blot.Cell viability assay,colony formation assay,wound healing assay and transwell assay in vitro and tumour xenograft studies in vivo were performed to assess the influence of miR-218 on the proliferation,invasion and migration of the BCa cell lines.Bioinformatics was used to predict the potential targets of miR-218.Luciferase reporter assay was used to demonstrate whether BMI-1 was a direct target of miR-218.Hematoxylin and eosin(HE)staining and immunohistochemical staining were performed to assess the proliferation of tumor tissues from nude mice.Results:The miR-218 expression level was frequently downregulated in BCa tissues compared with normal adjacent tissues.qRT-PCR showed microRNA-218 expression level of miR-218-transfected cell lines was 250 folds higher than that of control cell lines.The stable bladder cancer T24 and EJ cell lines overexpressing microRNA-218 was established.The overexpression of miR-218 significantly inhibited cell proliferation,migration and invasion in BCa cell lines via cell viability assay,colony formation assay,wound healing assay transwell assay in vitro and tumour xenograft studies in vivo.Dual luciferase reporter assay demonstrated that miR-218 mimics could reduce the fluorescence intensity in the wild-type BMI-1 3'-UTR vector(WT),while it had no effect on the mutant type BMI-1 3'-UTR vector(Mut).Luciferase reporter assay showed that BMI-1 was a direct target of miR-218.In addition,we found that miR-218 suppressed the expression of BMI-1 and its downstream target(PTEN)and participated in the regulation of phospho-AKT(pAKT).HE staining showed smaller nuclei in the miR-218-treated tumor compared with control group.Immunohistochemical staining identified that the level of nuclear proliferation antigen Ki-67 was downregulated in the miR-218-treated tumors compared with control group.Conclusion:Our study demonstrated that miR-218 was frequently downregulated in BCa tissues and functioned as a tumor suppressor by directly targeting BMI-1.Thus,the miR-218/BMI-1 axis may provide novel diagnostic and ther-apeutic strategies for the treatment of BCa.