Platelets prevent RBC leakage in inflamed vessels. (A) RBC leakage requires neutrophil transmigration in Geltrex mix vessels. Vessels were cultured against Geltrex mix (i) or collagen I (ii) for 1 week, and then stimulated overnight with 10 ng/mL TNF-α + 10 ng/mL IL-1β. Plts and neuts were isolated from human blood and resuspended in EGM-FCS. They were then perfused through the vessels for 1.5 hours. RBCs were added to the vessels for 30 minutes and then RBC leakage was imaged using an iPhone through a bright-field microscope. Dashed lines highlight the phase guides, the top of which marks the vessel wall. (B) Platelets prevent RBC leakage from Geltrex mix and collagen I vessels. RBC leakage was quantified in FIJI. (C) Single platelets adhere to inflamed vessels. Vessels were fixed, blocked, and stained with Hoechst-33342 (blue), α-CD41a (green), and α-CD31 (red). Z-stacks were taken on a Leica SP5 confocal microscope. N = 5 independent experiments per blood donor, n = 2 chips per condition. Mean ± SEM; ∗P < .05 and ∗∗P < .01. Scale bars are representative of 50 μm.