Figure 2.

pEVs inhibit angiogenesis in vivo. (a) 12-week-old male C57BL/6J mice were subcutaneously injected with either 5 µg of cEVs and pEVs, mixed with Matrigel Matrix supplemented with VEGF 100 ng/mL (450-32 Peprotech) and Heparin 50 units/mL. After 7 days, plugs were harvested. For haemoglobin quantification, plugs were processed by TissueLyser and the haemoglobin content was measured using Drabkin’s reagent kit 525 (Sigma-Aldrich). Each value was first normalized on the total plug protein quantity, measured by BCA assay, and then on the negative control (plugs with vehicle). (b, c) 1-day-old C57BL/6J mouse pups were intraperitoneally injected with a total of 10 μg of cEVs or pEVs, using PBS as control. Mice were sacrificed for retina collection. Dissected retina were stained with isolectin-b4 (green) and digital images were captured using inverted fluorescence confocal microscope. Analyses of the relative radial expansion and of the relative branching point were performed. All data are expressed as means ± SEM, normalized on control (mice treated with vehicle) (n = 8 mice/group). Ordinary one-way ANOVA; *P < 0.05.