Fig. 7

NETs activated the AKT/mTOR signaling pathway to promote angiogenesis. a The effect of NET-stimulated HUVECs on the injury factor vWF and pro-angiogenic factor ANGPT2 and the inhibitory effect of DNase-1 were observed via immunofluorescence. Magnification, × 20; scale bars: 50 μm; red, ANGPT2/vWF; green, CD31; blue, DAPI. b The average intensities of vWF and ANGPT2 were calculated using Image J software; all values are presented as means ± SDs. **p < 0.01; ***p < 0.001; ****p < 0.0001. c The tubule-forming ability of HUVECs with 0, 0.2, 0.4, and 0.8 μg/mL of NET stimulation and 25 ng/mL VEGF stimulation was detected via a tubule formation experiment. Magnification, × 20; scale bars: 50 μm. d The total length of vascular rings formed under different stimulation conditions was calculated using Image J software; all values are presented as means ± SDs. *p < 0.05; ****p < 0.0001. e The neovascularization ability was detected using the arterial ring germination test in nude mice with NET concentration gradients of 0, 0.2, 0.4, and 0.8 μg/mL and stimulation with 25 ng/mL VEGF. Magnification, × 20; scale bars: 50 μm. f Image J software was used to calculate the coverage area of vessels under different stimulation conditions; all values are presented as means ± SDs. ****p < 0.0001. g Western blotting was used to detect the activation and phosphorylation of the AKT/mTOR signaling pathway induced by NET-stimulated HUVECs. h Image J software was used to calculate the gray values of p-AKT/p-mTOR; all values are presented as means ± SDs. *p < 0.05; ****p < 0.0001. vWF, von Willebrand factor; ANGPT2, angiopoietin-2; SD, standard deviation