Fig. 3

The BBB permeability increases in the injured hemisphere at 6 and 24 h after neonatal HI. a Regional (hippocampus, cortex, striatum/thalamus) brain ¹⁴C-sucrose concentration ratios between left (injured) and right (uninjured) hemispheres at 6 (4 m, 5f), 24 (4 m, 4f) hours and 5 days (4 m, 5f) after neonatal HI. b Entire hemisphere brain ¹⁴C-sucrose concentration ratios between left (injured) and right (uninjured) hemispheres at various times between 6 h and 5 days after neonatal HI. Horizontal lines indicate mean value. * = p ≤ 0.05, ** = p ≤ 0.01, and *** = p ≤ 0.001 (paired t-test between hemispheres). c The vascular density of the brain is not altered at 5 days after hypoxia/ischemia in neonates. Vascular area defined as CLDN5⁺ area in brain sections (See Additional file 1 for more details). Vascular area of total area in cortex/hippocampus in both the left (injured) and right hemispheres of HI-animals (n = 5, 2 m, 4f) was similar to control animals (n = 5, 2 m, 4f). BBB-damage were confirmed by imaging brain sections from animals injected with the fluorescent dye Evans blue (EB). After transcardial perfusion, areas of EB-bound albumin in the brain parenchyma was present in the cortex of injured hemispheres (d) while the uninjured hemispheres only showed weak EB-fluorescence restricted to blood vessels in corresponding areas (e). Control animals injected with EB and not perfused showed strong EB-fluorescence in blood vessels and none in parenchyma (f). Extravasated EB encompassed 51.1 ± 8.4% of the total area in images from the injured hemisphere, significantly altered from the uninjured hemisphere (6.3 ± 1.9%, p = 0.0002) and controls (5.8 ± 1.6%, p = 0.0002) where the EB was restricted to blood vessels (g), cortical images were were taken from the region outlined in (h). n=3 per group. Scale-bars are 100 μm (d-f) and two mm (h). One-way ANOVA with Dunnett’s multiple comparison test. Data presented as mean ± SD. M male, f female. Bar graphs represent mean values.