MicroRNA‑204‑5p inhibits the osteogenic differentiation of ankylosing spondylitis fibroblasts by regulating the Notch2 signaling pathway.

Jianjun Zhao, Yanyan Zhang, Bo Liu

Molecular medicine reports 2020 Sep

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Ankylosing spondylitis (AS) is a chronic inflammatory systemic disease and is difficult to detect in the early stages. The present study aimed to investigate the role of microRNA (miR)‑204‑5p in osteogenic differentiation of AS fibroblasts. Bone morphogenetic protein 2 (BMP‑2) was used to induce osteogenic differentiation. Cells were divided into the following groups: AS group, AS + BMP‑2 group, AS + BMP‑2 + miR‑negative control group, AS + BMP‑2 + miR‑204‑5p mimics group and AS + BMP‑2 + miR‑204‑5p mimics + pcDNA‑Notch2 group. The expression levels of miR‑204‑5p, Notch2, runt‑related transcription factor 2 (RUNX2) and osteocalcin were detected via reverse transcription‑quantitative PCR analysis. The binding site between Notch2 and miR‑204‑5p was predicted using TargetScan software and verified via the dual‑luciferase reporter assay. Alkaline phosphatase (ALP) activity was assessed via the ALP assay, while the mineralized nodules area was determined via the Alizarin Red S staining assay. The results demonstrated that Notch2 is a target gene of miR‑204‑5p. Furthermore, treatment with BMP‑2 significantly decreased miR‑204‑5p expression, and significantly increased ALP activity, the mineralized nodules area and the expression levels of Notch2, RUNX2 and osteocalcin in ligament fibroblasts (all P<0.05). Conversely, transfection with miR‑204‑5p mimics significantly increased miR‑204‑5p expression, and significantly decreased ALP activity, the mineralized nodules area and the expression levels of Notch2, RUNX2 and osteocalcin in ligament fibroblasts (all P<0.05). Notably, transfection with pcDNA‑Notch2 significantly reversed the inhibitory effects induced by miR‑204‑5p mimics on the osteogenic differentiation of ligament fibroblasts (all P<0.05). Furthermore, miR‑204‑5p inhibited the osteogenic differentiation of ligament fibroblasts in patients with AS by targeting Notch2. Thus, miR‑204‑5p may negatively regulate Notch2 expression and may be a potential therapeutic target for AS. Collectively, the results of the present study provide a theoretical basis for the effective treatment of patients with AS.

Citation

Jianjun Zhao, Yanyan Zhang, Bo Liu. MicroRNA‑204‑5p inhibits the osteogenic differentiation of ankylosing spondylitis fibroblasts by regulating the Notch2 signaling pathway. Molecular medicine reports. 2020 Sep;22(3):2537-2544