Background: We aimed to investigate the biological effect of miR-497 in cigarette smoke extract (CSE)-damaged human bronchial epithelial (HBE) cells and the underlying molecular mechanism.
Methods: MiR-497 mimic was transfected into HBE cells to up-regulate miR-497 expression. Cigarette smoke extract (CSE, 20 μg/mL) was utilized to treat HBE cells to form the injury model. Cell proliferation and apoptosis were detected by CCK8 and flow cytometry assays. DDX3Y mRNA expression was determined by a quantitative reverse transcriptase-polymerase chain reaction. The interaction between miR-497 and DDX3Y was verified by a luciferase reporter assay. Protein expression levels were tested by western blotting.
Results: CSE treatment decreased miR-497 level in HBE cells. CSE exposure restrained cell proliferation, promoted cell apoptosis and enhanced the relative expression of TLR4 and p-NF-κB p65. DDX3Y was predicted as a target of miR-497. The mRNA and protein expression of DDX3Y was negatively modulated by miR-497 in CSE-injured HBE cells. Up-regulation of miR-497 by miR-497 mimic increased cell proliferation and reduced cell apoptosis in CSE-treated HBE cells, which were rescued by DDX3Y high expression in CSE-treated HBE cells. Consistently, Bcl-2 protein level was heightened, whereas Bax and actived caspase-3/9 protein levels were decreased by miR-497 mimic in CSE-stimulated HBE cells, which was reversed by DDX3Y over-expression in CSE-stimulated HBE cells. The relative expression of TLR4 and p-NF-κB p65 was decreased by miR-497 mimic, whereas they were rescued by DDX3Y over-expression in CSE-damaged HBE cells.
Conclusions: The results of the present study demonstrate that up-regulation of miR-497 exhibits a protective effect on CSE-damaged HBE cells, which might be achieved by targeting DDX3Y and regulating the TLR4/NF-κB pathway.
Keywords: DEAD-box helicase 3 Y-linked; Toll-like receptor 4/NF-ĸB pathway; chronic obstructive pulmonary disease; cigarette smoke extract; human bronchial epithelial cells; miR-497.
© 2019 John Wiley & Sons, Ltd.