miR-140-3p in exosomes derived from bone-marrow mesenchymal stem cells inhibits pyrotoisosis by targeting PTEN under high-glucose condition

Figure 1 Identification results of bone-marrow mesenchymal stem cell secreted exosome. A: Size distribution of bone-marrow mesenchymal stem cell secreted exosome (BMSC-Exo) detected by dynamic light scattering; B: The transmission electron microscopy images of the purified BMSC-Exo revealing the spheroid shape. Scale bar = 100 nm; C: Western blot analysis showing positive expression of exosomal markers CD9, CD63 and CD81 in BMSC-Exo.

Figure 2  Uptake of bone-marrow mesenchymal stem cell secreted exosomes by fibroblasts (n = 3).

Figure 3  Heatmap of differentially expressed microRNAs in exosomes derived from cells cultured under normal-glucose vs high-glucose conditions.

Figure 4 Results of the CCK-8 and scratch assays. A: Skin fibroblast proliferation in group B1 was inferior to that in control group A1. Group C1 displayed stronger cell division activity compared with group B1; B and C: Wound-healing rates in group B1 were lower than A1 group. Pyroptosis inhibitors enhanced fibroblast migration capacity in group C1. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001.

Figure 5 Western blot results. A-E: Compared with the control group, bone-marrow mesenchymal stem cell secreted exosomes transfected with the inhibitor of miR-140-3p could significantly increase the expression of pyroptosis-related proteins gasdermine-D-N-terminal (GSDMD-N) (A), cleaved caspase-1 (B), interleukin (IL)-1β (C), IL-18 (D), and NLRP3 (E) in skin fibroblasts under high-glucose conditions. Adding the pyroptosis pathway inhibitor AYC to the Hy + miR-140-3p inhibitor-Exo group could significantly decrease the expression of pyroptosis-related proteins GSDMD-N (A), cleaved caspase-1 (B), IL-1β (C), IL-18 (D), and NLRP3 (E); F: The expression of GSDMD-N, cleaved caspase-1, IL-1β, IL-18, and NLRP3 in cells with different treatments was detected by western blot. n = 6. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. GSDMD-N: Gasdermine-D-N-terminal; IL: Interleukin.

Figure 6 Polymerase chain reaction analysis of phosphatase and tensin homolog expression in each group. ^bP < 0.01. PTEN: Phosphatase and tensin homolog.

Figure 7 Western blot analysis of phosphatase and tensin homolog expression in each group. ^aP < 0.05, ^cP < 0.001. PTEN: Phosphatase and tensin homolog.

Figure 8 Dual-luciferase reporter assay results. ^cP < 0.001.

Figure 9 Relationship between the miR-140-3p/phosphatase and tensin homolog regulatory axis and pyroptosis-related proteins. ^aP < 0.05, ^bP < 0.01. GSDMD-N: Gasdermine-D-N-terminal; IL: Interleukin; PTEN: Phosphatase and tensin homolog.