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. 2023 Jul 1;29:85–97. doi: 10.1016/j.bioactmat.2023.06.011

Fig. 1.

Fig. 1

Antisenescence effects of ESC-EVs on late-passaged MEFs. (A) Schematic represention of EVs treating senescent MEFs. After 96 h of treatment with EVs, the 7th generation MEFs were used for subsequent detection. (B) Representative cellular morphology and SA-β-gal staining of P3 MEFs, P7 MEFs and P7 MEFs treated with MEF-EVs or ESC-EVs. Scale bar represents 100 μm. (C) Quantification of the percentage of SA-β-gal positive MEFs. (D) RT-qPCR analysis of senescence-related genes in P3 MEFs and P7 MEFs exposed to MEF-EVs and ESC-EVs, respectively. (E) Western blot analysis of p21 and p53 in P3 MEFs and P7 MEFs exposed to MEF-EVs and ESC-EVs, respectively. (F) Grayscale statistical analysis of p21 and p53 relative to H3. (G) Immunofluorescence analysis of p21 and p53 expression in P3 MEFs and P7 MEFs exposed to MEF-EVs or ESC-EVs. Scale bar represents 100 μm. Bottom panel: Quantification of the percentage of p21 and p53 positive MEFs. (H) Immunofluorescence analysis of Ki67 and γ-H2AX expression in P3 MEFs and P7 MEFs exposed to MEF-EVs or ESC-EVs. Scale bar represents 100 μm. Bottom panel: Quantification of the percentage of Ki67-and γ-H2AX-positive MEFs. (I) Cell cycle of P3 MEFs and P7 MEFs exposed to MEF-EVs or ESC-EVs were presented by G0/G1, S and G2/M phases. (J) Quantification of the ratio of ROS-positive cells in each group to the negative control group. Data are presented as mean ± SEM. n = 3. Student's t-test: ns, not significance; *p < 0.05 and **p < 0.01.