Fig. 4

Overexpression of miR-204-5p suppresses osteogenic differentiation of SCAPs. a Western blot assays were performed to analyze the protein expression levels of ALP, RUNX2, and OSX in the miR-204-5p group than mimics NC group. GAPDH was the internal control. b Grayscale analyses. c qRT-PCR showed lower levels of ALPL, RUNX2, and OSX in the mimics group. d, g Images of ALP staining in the miR-204-5p group. SCAPs were cultured in OM for 5 days. Histograms showed ALP activity by spectrophotometry (bar = 200 μm) e, h After cell culture in OM for 14 days, Alizarin Red S staining, and CPC assay showed that miR-204-5p mimics suppresses mineralization of SCAPs (scale bar = 200 μm). f Transfection efficacy of miR-204-5p mimics and inhibitor. i Immunofluorescence staining showed negative expressions of ALP and RUNX2 in the miR-204-5p mimics group (scale bar = 50 μm. Quantification was done by ImageJ. *P < 0.05, **P < 0.01, ***P < 0.001