Fig. 3

Stimulated angiogenesis mediated by ADSC-NVs in vitro. The dose-effect of ADSC-NVs and ADSC-EVs on cell migration (C) and tube formation (D) abilities of HUVECs was confirmed. Considering the possible side effects and enormous economic cost of high concentrations [28], different doses of ADSC-EVs and ADSC-NVs (ranging from 5 to 80 μg/mL) were assessed. Transwell assay was conducted following the shown schematic diagram (A). Briefly, HUVECs on the upper side of the transwell membrane were treated with different dose of ADSC-NVs or ADSC-EVs (0, 5, 20 and 80 μg/mL) in the lower side for 24 h. The cells migrated to the bottom of the membrane were fixed with 4% paraformaldehyde and stained with crystal violet. C Representative images of transwell assay with ADSC-EVs or ADSC-NVs treatment. Scale bar: 10 μm. B Quantitative analysis of the migrated cells (n = 4 per group). D Formation of tube-like structures by HUVECs-GFP after incubation with different doses of ADSC-EVs or ADSC-NVs (0, 5, 20, and 80 μg/mL) for 6 h. Scale bar: 100 μm. E Quantitative analysis of tube-like structures formation (n = 3 per group). F mRNA expression of angiopoietin-1 (Ang-1) and vascular endothelial growth factor (VEGF) in HUVECs treated with ADSC-NVs or ADSC-EVs was confirmed using qRT-PCR (n = 3 per group). G Expression levels of Ang-1 and VEGF proteins in HUVECs after ADSC-NVs or ADSC-EVs stimulation. ns, no significant difference, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001