Overexpression of MiR-140-5p can Significantly Improve IL-1β-Induced Chondrocytes Cell Viability and Inflammatory Response

Abstract

Objective: To investigate the biological function of the miR-140-5p/Nrf2 (nuclear factor erythroid 2-Related factor 2) axis in the development and progression of osteoarthritis. Methods: The optimal concentration of Interleukin-1β (IL-1β)-induced mouse chondrocytes ATDC-5 cells was selected by CCK-8 (cell counting kit-8), subsequent to the construction of an in vitro osteoarthritis model. Quantitative real-time polymerase chain reaction (RT-qPCR) was applied to test the transfection efficiency of ATDC-5 cells and miR-140-5p mimics. Additionally, cell apoptosis and cycle were determined by flow cytometry, cell viability was observed through CCK-8 assay, and Heme Oxygenase-1 (HO-1), Nrf2, B-cell lymphoma-2-Associated X (BAX), and B-cell lymphoma-2 (Bcl-2) protein expression levels were analyzed via western blotting. IL-6, TNF (tumor necrosis factor)-α and MDA (malonyldialdehyde) were studied by conducting ELISA (enzyme-linked immunosorbent assay);And the targeting association of Nrf2 and miR-140-5p, was by dual-luciferase reporter assay analysis. Results: The outcomes suggested that IL-1β (10 ng/mL) notably inhibited ATDC-5 cell viability. Relative to controls, Nrf2 protein expression markedly rose and miR-140-5p levels remarkably declined in ATDC-5 cells following IL-1β treatment. Dual-luciferase reporter test indicated that Nrf2 served as a miR-140-5p target gene. In addition, miR-140-5p overexpression greatly promoted IL-1β-induced ATDC-5 cell viability and inhibited ATDC-5 cell cycle arrest and apoptosis. Moreover, miR-140-5p overexpression significantly decreased Nrf2, BAX and HO-1 levels while increased Bcl-2 in IL-1β-induced ATDC-5 cells. Moreover, miR-140-5p overexpressed in ATDC-5 cells down-regulated intracellular inflammatory cytokines—MDA, IL-6 and TNF-α levels. Conclusions: Altogether, miR-140-5p may alleviate osteoarthritis by inhibiting inflammatory response, chondrocyte apoptosis, cell cycle arrest, and oxidative stress.