miR-495 andmiR-5688 are down-regulated in non-small cell lung cancer under hypoxia to maintain interleukin-11 expression

Background:Hypoxia is a hallmark of cancer and is associated with poor prognosis.However,the molecular mechanism by which hypoxia promotes tumor progression remains unclear.MicroRNAs dysregulation has been shown to play a critical role in the tumor and tumor microenvironment.Here,we investigated the roles ofmiR-495 and miR-5688 in human non-small cell lung cancer(NSCLC)and their underlying mechanism.Methods:The expression levels of miR-495 and miR-5688 in human NSCLC tissue specimens were measured by quantitative real-time polymerase chain reaction(qRT-PCR).Deferoxamine(DFO)was used to determine whether the regulation of miR-495 and miR-5688 under hypoxia was dependent on hypoxia-inducible factor 1-alpha(HIF-1α).Furthermore,the functions of miR-495 and miR-5688 in tumor progression were evaluated using colony formation,3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2Htetrazolium(MTS),wound healing,transwell assays,and xenograft model.Two algorithms,PicTAR and Targetscan,were used to predict the target gene of these two miRNAs,and dual-luciferase reporter assay was conducted to confirm the target.The unpaired two-tailed t test,Pearson correlation analysis,and Fisher’s exact probability test were performed for statistical analyses.Results:Two miRNAs,miR-495 and miR-5688,were found to participate in NSCLC progression under hypoxia.They were down-regulated in NSCLC tissues compared with normal tissues.We determined that hypoxia led to the down-regulation of miR-495 and miR-5688 in NSCLC cells,which was independent of HIF-1αand cellular metabolic energy.In addition,miR-495 and miR-5688 suppressed cell proliferation,migration,and invasion in vitro.The NSCLC xenograft model showed that miR-495 and miR-5688 inhibited tumor formation in vivo.Interestingly,we found that miR-495 and miR-5688 had the same target,interleukin-11(IL-11).Recombinant human IL-11 counteracted the effects of miR-495 and miR-5688 on NSCLC cells,suggesting that miR-495 and miR-5688 executed their tumor suppressive role by repressing IL-11 expression.Conclusion:We found that hypoxia down-regulated the expression levels of miR-495 and miR-5688 in NSCLCto enhance IL-11 expression and tumor progression,indicating that the miR-495/miR-5688/IL-11 axismay serve as a therapeutic target and potential biomarker for NSCLC.