Figure 3. 3D BMSCs coculture activates ZMPSTE24 signaling. (A–C) Immunofluorescence staining of ZMPSTE24 between young (3 weeks) and old (24 months) rats, scale bar, 50um. Fluorescence intensities were determined by using Image J software. (B–D) Protein expression of P16, P21 and ZMPSTE24 were visualized by western blot and quantified by Image J. (E–F) Protein expression of ZMPSTE24 during coculture was visualized by western blot and quantified by Image J. (G) Immunofluorescence staining of ZMPSTE24 and MMP9 in NP cells between Sene, Sene +NPCs and Sene +BMSCs groups. n=5. Scale bar=50um. (H) Heat map representation of SASP (senescence-associated secretory phenotype) genes between CTL, Sene +NPCs and Sene +BMSCs groups. n=3 biological replicates. Values represent means±S.D. Significant differences between different groups are indicated as *P < 0.05, **P < 0.01, ***P < 0.001.