Fig. 4

miR-325-3p loss in chondrocytes precipitates senescent states and miR-325-3p overexpression alleviates mechanical stress-induced senescence in vitro. A Representative p21 immunofluorescence staining and SA-β-Gal images in chondrocytes treated with miR-325-3p control or inhibitor. Red arrows represent SA-β-Gal⁺ cells. Scale bar: 20 μm in the left SA β-Gal staining panel and 50 μm in the right IF panel. B, C Quantification of p21 immunofluorescence and SA-β-Gal staining in A. n = 6 per time point. D Quantitative PCR analysis of SASP-related genes (PAI-1, IL-6, TGF-β) in chondrocytes treated with miR-325-3p inhibitor or NC-inhibitor. n = 3 per group. E, F Representative western blot images and quantification in chondrocytes treated with miR-325-3p inhibitor or NC-inhibitor. n = 3 per group. G Representative p21 immunofluorescence staining and SA-β-Gal images in CTS (20% elongation, 24 h)-treated chondrocytes when administrated with miR-325-3p mimic or NC-mimic. Red arrows represent SA-β-Gal.⁺ cells. Scale bar: 20 μm in the left SA β-Gal staining panel and 50 μm in the right IF panel. H, I Quantification of p21 immunofluorescence and SA-β-Gal staining in G. n = 6 per time point. J Quantitative PCR analysis of SASP-related genes (PAI-1, IL-6, TGF-β) in CTS (20% elongation, 24 h)-treated chondrocytes treated with miR-325-3p mimic or NC-mimic. n = 3 per group. K, L Representative western blot images and quantification in CTS (20% elongation, 24 h)-treated chondrocytes treated with miR-325-3p mimic or NC-mimic. n = 3 per group. All data are shown as the mean ± standard deviation (SD). *P < 0.05, **P < 0.01