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

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.