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

Abstract

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 of miR-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)-2H-tetrazolium (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-1α and 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 NSCLC to enhance IL-11 expression and tumor progression, indicating that the miR-495/miR-5688/IL-11 axis may serve as a therapeutic target and potential biomarker for NSCLC.