Key WordsInflammatory injury • MicroRNA-146a • C-X-C chemokine receptor type 4 • PI3K/AKT • Wnt/β-catenin Abstract Background/Aims: Osteoarthritis (OA) as a degenerative disease is a major problem in ageing populations. To better understand the molecular mechanisms in the pathogenesis of OA, this study explored the role of microRNA (miR)-146a in the articular chondrocytes. Methods: The articular chondrocyte line ATDC5 was used to simulate inflammatory injury by LPS administration in vitro. Cell viability, apoptosis, mRNA expressions and productions of inflammatory factors were assessed, respectively. Mir-146a and Cxcr4 mRNA expressions were measured by qRT-PCR. Targeting effect of miR-146a on Cxcr4 3'UTR was assessed by luciferase activity analysis. Protein expression levels of CXCR4 and main factors in PI3K/AKT, Wnt/β-catenin signal pathways were measured by western blotting. Results: LPS exposure suppressed cell viability, prompted apoptosis of ATDC5 cells, and stimulated expression and release of inflammatory factors. MiR-146a was upregulated in LPS-induced cells. Overexpression of miR146a further aggravated LPS-induced inflammatory injury, while it was reduced after miR146a was knocked down. CXCR4 expression was negatively regulated by miR-146a. CXCR4 was a direct target of miR-146a and thus involved in regulatory effect of miR-146a on the injured chondrocytes, which was also related with phosphorylation levels of PI3K/AKT and expressions of Wnt/β-catenin signal factors. Conclusion: miR-146a promoted inflammatory response of articular chondrocytes via targeting CXCR4 and suppressing CXCR4 expression. Overexpression of CXCR4 could attenuate the inflammatory injury. Our findings provided novel evidence which might be useful for further studies exploring therapeutic approaches for OA via targeting miR-146a.

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Background/Aims: Several long non-coding RNAs (lncRNAs) play vital roles in osteoarthritis (OA), whereas the role of lncRNA RP11-445H22.4 in OA remains unclear. The study aimed to investigate the effect of lncRNA RP11-445H22.4 on lipopolysaccharide (LPS)-induced cell viability, apoptosis and inflammatory injury of OA. Methods: The expression of RP11-445H22.4, miR-301a and CXCR4 in human cartilage ATDC5 cells were altered by transfection, and then cells were exposed to 5 µg/ml LPS for 12 h. Then cell viability, apoptosis, apoptosis-related factors and inflammatory cytokines were analyzed by CCK-8, flow cytometry, western blot, RT-qPCR and ELISA, respectively. Dual-luciferase reporter assay was performed to assess the binging sites of RP11-445H22.4 and miR-301a. The signal pathways of NF-κB and MAPK/ ERK were determined by western blot. Results: LPS reduced cell viability, increased apoptosis and stimulated release of IL-1β, IL-6, IL-8 and TNF-α. However, RP11-445H22.4 inhibition significantly rescued LPS-induced injuries by promoting cell viability, suppressing apoptosis and inflammatory cytokines secretions in ATDC5 cells. In addition, miR-301a directly bound to RP11-445H22.4, and suppression of miR-301a inversed the effects of RP11-445H22.4 inhibition. Furthermore, CXCR4 was a direct target of miR-301a, and CXCR4 silencing increased cell viability, decreased apoptosis and inflammatory cytokines secretions in LPS-treated ATDC5 cells. Besides, we found that CXCR4 silencing blocked LPS-activated NF-κB and MAPK/ERK pathways. Conclusions: The study indicated that lncRNA RP11-445H22.4-miR-301a-CXCR4 axis played an important role in cartilage ATDC5 cells and provided a theoretical basis of lncRNA RP11-445H22.4 in OA.

Background: The anti-inflammatory function of microRNA-124 (miR-124) has been a matter of extensive studies in the last few years. Although LINC00305 regulates biological activities by acting as a miR sponge, it is still unexplored whether LINC00305 is involved in inflammation by regulating miR-124. Methods: Cell viability and apoptosis were evaluated with commercial kits, cell counting kit-8 (CCK-8) and Annexin V-fluorescein isothiocyanate (FITC) kit, respectively. LINC00305, miR-124 and mRNA levels were quantified by quantitative reverse transcription PCR (qRT-PCR). Protein level was assessed with Western blot assay and enzyme-linked immunosorbent assay (ELISA). Results: The expression of LINC00305 was up-regulated by lipopolysaccharide (LPS). LINC00305 overexpression further suppressed the cell viability, promoted apoptosis and induced inflammation in LPStreated ATDC5 cells while its silence enhanced the cell viability, inhibited apoptosis and ameliorated inflammation. miR-124 was negatively regulated by LINC00305 and its overexpression abolished the effects of LINC00305 in the presence of LPS. LINC00305 further triggered the Notch/nuclear factor kappa B (NF-jB) signalling pathway in LPS-treated ATDC5 cells and this activation was abrogated when ATDC5 cells overexpressed miR-124. Conclusion: LINC00305 might emerge as a novel suppressor for miR-124. LINC00305-caused miR-124 silence compromises ATDC5 cell viability and ultimately results in inflammatory insults by activating Notch/NF-jB pathway.

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