Downregulation of MicroRNA-495 Alleviates IL-1ß Responses among Chondrocytes by Preventing SOX9 Reduction.

PURPOSE: Our previous work demonstrated that miRNA-495 targets SOX9 to inhibit chondrogenesis of mesenchymal stem cells. In this study, we aimed to investigate whether miRNA-495-mediated SOX9 regulation could be a novel therapeutic target for osteoarthritis (OA) using an in vitro cell culture model. MATERIALS AND METHODS: An in vitro model mimicking the OA environment was established using TC28a2 normal human chondrocyte cells. Interleukin-1ß (IL-1ß, 10 ng/mL) was utilized to induce inflammation-related changes in TC28a2 cells. Safranin O staining and glycosaminoglycan assay were used to detect changes in proteoglycans among TC28a2 cells. Expression levels of COX-2, ADAMTS5, MMP13, SOX9, CCL4, and COL2A1 were examined by qRT-PCR and/or Western blotting. Immunohistochemistry was performed to detect SOX9 and CCL4 proteins in human cartilage tissues obtained from patients with OA. RESULTS: miRNA-495 was upregulated in IL-1ß-treated TC28a2 cells and chondrocytes from damaged cartilage tissues of patients with OA. Anti-miR-495 abolished the effect of IL-1ß in TC28a2 cells and rescued the protein levels of SOX9 and COL2A1, which were reduced by IL-1ß. SOX9 was downregulated in the damaged cartilage tissues of patients with OA, and knockdown of SOX9 abolished the effect of anti-miR-495 on IL-1ß-treated TC28a2 cells. CONCLUSION: We demonstrated that inhibition of miRNA-495 alleviates IL-1ß-induced inflammatory responses in chondrocytes by rescuing SOX9 expression. Accordingly, miRNA-495 could be a potential novel target for OA therapy, and the application of anti-miR-495 to chondrocytes could be a therapeutic strategy for treating OA.

Ubiquitin-specific protease 53 promotes osteogenic differentiation of human bone marrow-derived mesenchymal stem cells.

The ubiquitin protease pathway plays important role in human bone marrow-derived mesenchymal stem cell (hBMSC) differentiation, including osteogenesis. However, the function of deubiquitinating enzymes in osteogenic differentiation of hBMSCs remains poorly understood. In this study, we aimed to investigate the role of ubiquitin-specific protease 53 (USP53) in the osteogenic differentiation of hBMSCs. Based on re-analysis of the Gene Expression Omnibus database, USP53 was selected as a positive regulator of osteogenic differentiation in hBMSCs. Overexpression of USP53 by lentivirus enhanced osteogenesis in hBMSCs, whereas knockdown of USP53 by lentivirus inhibited osteogenesis in hBMSCs. In addition, USP53 overexpression increased the level of active ß-catenin and enhanced the osteogenic differentiation of hBMSCs. This effect was reversed by the Wnt/ß-catenin inhibitor DKK1. Mass spectrometry showed that USP53 interacted with F-box only protein 31 (FBXO31) to promote proteasomal degradation of ß-catenin. Inhibition of the osteogenic differentiation of hBMSCs by FBXO31 was partially rescued by USP53 overexpression. Animal studies showed that hBMSCs with USP53 overexpression significantly promoted bone regeneration in mice with calvarial defects. These results suggested that USP53 may be a target for gene therapy for bone regeneration.