Fig. 4

Effects of BMSC-Exos and CC-Exos on the proliferation and differentiation of CPCs. a Immunofluorescence staining revealed that there was no difference between the two kinds of exosomes internalized by CPCs. Exosomes are red (CM-Dil) and overlaid on cultures of proliferating CPCs for 12 h. Scale bar = 20 μm. b Immunofluorescence of Ki67 in CPCs incubated with BMSC-Exos and CC-Exos (10 μg/mL and 30 μg/mL) for 12 h. CPCs were stained for Ki67 and DAPI (nucleus) and considered positive when Ki67 overlapped the nucleus. In these images, Ki67 are green, exosomes are red (CM-Dil), and nuclei are blue (DAPI). Scale bar = 50 μm. c Proliferation of CPCs after BMSC-Exos and CC-Exos stimulation and the negative control for 1, 3, 5, and 7 days. d Gene analysis for chondrogenesis and angiogenesis of CPCs stimulated by different exosomes. qRT-PCR results demonstrated that the CC-Exos group showed the highest expression of cartilage-associated genes, such as COL II and SOX-9, and the angiogenesis-associated genes, such as VEGF and SDF-1. e Western blot analysis of TGF-β, P-SMAD2/3, T-SMAD2/3, COL II, and SOX-9 secreted by CPCs exposed to exosomes for 48 h. f Western blot analysis of angiogenesis-associated VEGF and SDF-1 secreted by CPCs exposed to exosomes for 48 h. Data are shown as the mean ± standard deviation for n = 3. *P < 0.05, compared to negative control (NC), ^#P < 0.05, CC-Exos group compared to BMSC-Exos group