MiR-26b regulates cartilage differentiation of bone marrow mesenchymal stem cells in rats through the Wnt/β-catenin signaling pathway

T. Huang, Y. Zhou, J. Wang, Y. Cao, D.-H. Hang

Department of Orthopaedics, Baoshan Branch of Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, P.R. China. llrr197908@163.com

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• Stem Cells

OBJECTIVE: To investigate the influence of micro-ribonucleic acid (miR)-26b on the cartilage differentiation of mesenchymal stem cells (MSCs) in rats and its mechanism. This study aims to provide references for the clinical treatment of orthopedic diseases, such as osteoarthritis.

MATERIALS AND METHODS: MSCs were isolated from rat bone marrow, followed by identification of their immunological manifestation and multi-lineage differentiation potential. In addition, miR-26b small-interfering RNA (siRNA) was transfected into rat MSCs for evaluating its regulatory effect on MSCs differentiation. The predicted target gene Wnt was detected via Luciferase reporter gene assay and further verified via Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and Western blotting.

RESULTS: The expression level of miR-26b was significantly down-regulated during the in vitro cartilage differentiation of rat MSCs. Transfection of miR-26b siRNA enhanced in vitro cartilage differentiation of MSCs, and upregulated expressions of chondrogenesis-related marker molecules, including Collagen II and Aggrecan. Alcian blue staining further revealed that the differentiation of rat MSCs was promoted after transfection of miR-26b siRNA. According to further studies, the Wnt/β-catenin signaling pathway was significantly activated during the differentiation of MSCs, and its expression was negatively regulated by miR-26b. The results of the Luciferase reporter gene assay showed that miR-26b could directly inhibit the 3’untranslated region (3’UTR) of Wnt in a targeted manner.

CONCLUSIONS: MiR-26b plays an inhibitory role in the in vitro cartilage differentiation of rat MSCs by inhibiting Wnt expression.

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