Characterization of BMSCs. (A) Morphology of BMSCs derived from 2 young healthy donors (young BMSCs; passage 2; donor ages: 19 and 20 years old) and 2 older healthy donors (older BMSCs; passage 2; donor age: 68 and 72 years) by phase-contrast inverted microscopy. Scale bar, 50 µm. (B) BMSC growth measured after 24, 48, 72, 96, and 120 h in culture. Cell proliferation of young BMSCs (pink line) and older BMSCs (blue) was evaluated. Each value represents the mean ± SD (n = 6). (C-D) Representative images of SA-β-gal staining (arrows indicate positive cells) and quantification of cell senescence. Data represent the mean ± SD. *P < .01 in comparison with young BMSCs using a Student t test. Scale bar, 50 µm. (E-F) Telomere lengths in young and older BMSCs determined by Southern blotting of TRFs. Data represent the mean ± SD. *P < .05 in comparison with young BMSCs using a Student t test. (G) Transmission electron micrographs of exosomes derived from young and older BMSCs. Scale bar, 50 nm. (H) Western blots of exosomal lysates (CD63 and CD81 are common exosomal markers) derived from young and older BMSCs. Exosomes were isolated from 5 mL of culture medium collected from BMSCs (4 × 10⁴ cells/cm²) grown for 48 h. Equal volumes of exosomal lysate (30 µL) were loaded in the lanes of gels. (I) Nanoparticle concentration and size distribution of exosomes derived from young BMSCs (pink line) and older BMSCs (blue line).