Figures & data
Figure 1. Characterization of Exos: (A) Morphology of Exos observed by TEM. The scale bar is 100 nm. (B) Particle size and distribution analysis of Exos. (C) Western blotting of Exos surface markers.
![Figure 1. Characterization of Exos: (A) Morphology of Exos observed by TEM. The scale bar is 100 nm. (B) Particle size and distribution analysis of Exos. (C) Western blotting of Exos surface markers.](/cms/asset/dee15097-c2e2-4e8a-911e-6ac2c24b042f/ianb_a_1669617_f0001_c.jpg)
Figure 2. When low Exos concentrations (50 μg/mL) (Exos low) and high Exos concentrations (500 μg/mL) (Exos high) and the combination of high Exos (500 μg/mL) and LPS (100 ng/mL) concentrations (Exos high + LPS) were used to treat HG-HUVECs, Exos induced pro-angiogenic effects. untreated HG-HUVECs was used as the control group. (A) Exospromoted HG-HUVEC migration, as analyzed by scratch wound assay; this effect was reduced by Exos + LPS. (B) The migrating images of HG-HUVECs receiving different treatments were further confirmed by the transwell assay. Representative images of HG-HUVEC tube formation in different treatment groups. (C) Quantitative analysis of the scratch wound assay, transwell assay, and the tube-formation assay. The data are represented as the means ± SD (n = 3), #p<.05, compared untreated HG-HUVECs (control) with high Exos levels (500 μg/mL).*p < .05. (D) Cell viability was analyzed by CCK-8 assay with different treatments. The data are represented as the means ± SD (n = 3), *p < .05.
![Figure 2. When low Exos concentrations (50 μg/mL) (Exos low) and high Exos concentrations (500 μg/mL) (Exos high) and the combination of high Exos (500 μg/mL) and LPS (100 ng/mL) concentrations (Exos high + LPS) were used to treat HG-HUVECs, Exos induced pro-angiogenic effects. untreated HG-HUVECs was used as the control group. (A) Exospromoted HG-HUVEC migration, as analyzed by scratch wound assay; this effect was reduced by Exos + LPS. (B) The migrating images of HG-HUVECs receiving different treatments were further confirmed by the transwell assay. Representative images of HG-HUVEC tube formation in different treatment groups. (C) Quantitative analysis of the scratch wound assay, transwell assay, and the tube-formation assay. The data are represented as the means ± SD (n = 3), #p<.05, compared untreated HG-HUVECs (control) with high Exos levels (500 μg/mL).*p < .05. (D) Cell viability was analyzed by CCK-8 assay with different treatments. The data are represented as the means ± SD (n = 3), *p < .05.](/cms/asset/d54cf933-daf9-4777-9eca-5d2c78e3c1ce/ianb_a_1669617_f0002_c.jpg)
Figure 3. Detection of TNF-α, IL-6, AKT, P-AKT, and VEGF levels by Western blotting. The data are represented as the means ± SD (n = 3), #p<.05, compared untreated HG-HUVECs (control) with high Exos levels (500 μg/mL).*p < .05.
![Figure 3. Detection of TNF-α, IL-6, AKT, P-AKT, and VEGF levels by Western blotting. The data are represented as the means ± SD (n = 3), #p<.05, compared untreated HG-HUVECs (control) with high Exos levels (500 μg/mL).*p < .05.](/cms/asset/4e943290-f05b-4b25-aaff-c5bb6172383b/ianb_a_1669617_f0003_c.jpg)
Figure 4. (A) Representative images of full-thickness skin defects in a diabetic rat model untreated (PBS) or treated with low Exos (100 μg/mL), high Exos (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) at 0 days, 7 days, 14 days, and 21 days after operation. The scale bar in all is 5 mm. (B) Transmitted light images of H&E-stained sections of the untreated defects (PBS) and defects treated with low Exos, high Exos, or high Exos + LPS at day 7 after operation. (C) The wound size reduction in wounds receiving different treatments based on representative images of full-thickness skin defects in a diabetic rat model. The data are represented as the means ± SD (n = 3), #p<.05, compared PBS group with high Exos levels (1 mg/mL), *p < .05. (D) Determination of wound contraction rate based on transmitted light images of H&E-stained sections. The data are represented as the means ± SD (n = 3), *p < .05.
![Figure 4. (A) Representative images of full-thickness skin defects in a diabetic rat model untreated (PBS) or treated with low Exos (100 μg/mL), high Exos (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) at 0 days, 7 days, 14 days, and 21 days after operation. The scale bar in all Figure 4(A) is 5 mm. (B) Transmitted light images of H&E-stained sections of the untreated defects (PBS) and defects treated with low Exos, high Exos, or high Exos + LPS at day 7 after operation. (C) The wound size reduction in wounds receiving different treatments based on representative images of full-thickness skin defects in a diabetic rat model. The data are represented as the means ± SD (n = 3), #p<.05, compared PBS group with high Exos levels (1 mg/mL), *p < .05. (D) Determination of wound contraction rate based on transmitted light images of H&E-stained sections. The data are represented as the means ± SD (n = 3), *p < .05.](/cms/asset/3a8e8dfa-dba9-4ae4-b43b-68ab5df63fc1/ianb_a_1669617_f0004_c.jpg)
Figure 5. H&E staining of tissue sections treated with PBS, low Exos concentrations (100 μg/mL), high Exos concentrations (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) concentrations on days 7 and 14. The scale bar in figure is 50 μm.
![Figure 5. H&E staining of tissue sections treated with PBS, low Exos concentrations (100 μg/mL), high Exos concentrations (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) concentrations on days 7 and 14. The scale bar in figure is 50 μm.](/cms/asset/a1920084-ae40-4f49-9c28-41ad4012ea4d/ianb_a_1669617_f0005_c.jpg)
Figure 6. Masson’s trichrome staining of wound sections treated with PBS, low Exos concentrations (100 μg/mL), high Exos concentrations (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) concentrations at days 7 and 14. The data are presented as the means ± SD (n = 3); *p < .05. The scale bar in all figure is 200 μm. The relative pigmentation area (%) was determined by calculating the ratio of area of the blue stain (collagen fibers) to the whole selected area.
![Figure 6. Masson’s trichrome staining of wound sections treated with PBS, low Exos concentrations (100 μg/mL), high Exos concentrations (1 mg/mL), or high Exos (1 mg/mL) + LPS (10 μg/mL) concentrations at days 7 and 14. The data are presented as the means ± SD (n = 3); *p < .05. The scale bar in all figure is 200 μm. The relative pigmentation area (%) was determined by calculating the ratio of area of the blue stain (collagen fibers) to the whole selected area.](/cms/asset/0c703c44-cc05-45df-aa47-9d730934f210/ianb_a_1669617_f0006_c.jpg)
Figure 7. (A) Immunofluorescence staining of CD31. Newly formed blood vessels were identified by positive CD31 staining at postoperative day 7. The scale bar in figure 7 A is 50 μm. (B) Quantitative analysis of the number of blood vessels. The data are represented as the means ± SD (n = 3), *p < .05.
![Figure 7. (A) Immunofluorescence staining of CD31. Newly formed blood vessels were identified by positive CD31 staining at postoperative day 7. The scale bar in figure 7 A is 50 μm. (B) Quantitative analysis of the number of blood vessels. The data are represented as the means ± SD (n = 3), *p < .05.](/cms/asset/7298ad35-4103-47af-908e-02b8839c1f04/ianb_a_1669617_f0007_c.jpg)
Figure 8. Immunohistochemistry analysis of IL-6 expression and TNF-α expression in diabetic wound sites treated with PBS, low concentrations of Exos, high concentrations of Exos, and high Exos + LPS concentrations at days 7 and 14; quantitative analysis of IL-6 expression and TNF-α expression. The data are presented as the means ± SD (n = 3); *p < .05.
![Figure 8. Immunohistochemistry analysis of IL-6 expression and TNF-α expression in diabetic wound sites treated with PBS, low concentrations of Exos, high concentrations of Exos, and high Exos + LPS concentrations at days 7 and 14; quantitative analysis of IL-6 expression and TNF-α expression. The data are presented as the means ± SD (n = 3); *p < .05.](/cms/asset/eb3521cf-ec31-4896-98a3-f29102be4296/ianb_a_1669617_f0008_c.jpg)