miR-31-5p promotes NSCLC cell aggressiveness and disrupts the in vitro blood-brain barrier by negatively regulating the QKI/TUG1 pathway.

OBJECTIVE: To investigate the role of miR-31-5p in non-small cell lung cancer (NSCLC) cells and its function in an in vitro blood-brain barrier (BBB) model, and to elucidate the underlying regulatory mechanisms. METHODS: RT-qPCR was used to determine the relative expression levels of miR-31-5p, QKI mRNA, and TUG1 in serum samples obtained from the healthy control group and patients with NSCLC brain metastasis. Lentiviruses encoding miR-31-5p mimic, miR-31-5p inhibitor, and the corresponding negative control were transduced into A549 and NCI-H1975 cells. The effects of miR-31-5p on A549 and NCI-H1975 cells were assessed using RT-qPCR, cell cycle assay kits, Annexin V-FITC/PI apoptosis detection kits, wound healing assays, and Transwell assays. To explore the role of miR-31-5p/QKI/TUG1 axis in the blood-brain barrier (BBB) in vitro, bEnd.3 cells transfected with pcDNA-QKI and pcDNA-TUG1 were co-cultured with A549 or NCI-H1975 cells. RT-qPCR, Western blot, flow cytometry, transendothelial electrical resistance (TEER) measurement, and FITC-dextran assays were used to investigate the regulatory effect of miR-31-5p on the QKI/TUG1 axis and its subsequent impact on an in vitro BBB model. Dual-luciferase reporter assay and RNA immunoprecipitation assays confirmed the direct interactions among QKI, miR-31-5p, and lncRNA TUG1. RESULTS: Overexpression of miR-31-5p promoted cell cycle progression, invasion, and migration in A549 and NCI-H1975 cells, while it inhibited apoptosis. Furthermore, overexpression of miR-31-5p inhibited the expression of tight junction proteins in bEnd.3 cells, reduced the TEER value, and increased cell permeability. Knockdown of miR-31-5p produced effects opposite to those of miR-31-5p overexpression. Mechanistic studies revealed that miR-31-5p targeted and negatively regulated QKI, and that QKI negatively regulated TUG1 expression. Overexpression of QKI suppressed TUG1 expression, reduced apoptosis in bEnd.3 cells, and promoted TJ protein expression. Overexpression of TUG1 reversed the effects of QKI overexpression in bEnd.3 cells. In NSCLC brain metastasis patients, miR-31-5p and TUG1 levels were upregulated, whereas QKI mRNA level was downregulated. CONCLUSION: Knockdown of miR-31-5p suppresses cell cycle progression, migration, and invasion, induces apoptosis in A549 and NCI-H1975 cells, and attenuates bEnd.3 cell-mediated BBB injury in vitro through the QKI/TUG1 axis.