Figure 3.

Extravasated Alb-mSc-ST is captured by ECS-facing SC in astrocytes through ST–SC binding.

(a) Alb-mSc-ST mice injected with AAVs for astrocyte-targeted expression of ECS-facing SC are used for detecting BBB leakage, as extravasated Alb-mSc-ST will be bound by available ECS-SC at astrocytic endfeet.

(b) HEK293T cells expressing either ECS-SC or degradable ECS-dSC were exposed to Alb-mSc-ST plasma and imaged 2h and 28 h post-plasma addition (top). Fluorescence intensity at 2h and 28h (bottom, left) and their ratio (bottom, right) for ECS-SC- and ECS-dSC-expressing cells, showing faster turnover of ECS-dSC.

(c) Experiment timeline.

(d) Example images of vessels before and after laser irradiation in naïve Alb-mSc-ST mice and Alb-mSc-ST mice expressing ECS-SC or ECS-dSC in astrocytes. Scale bar, 10 μm.

(e) Example images of laser-irradiated and contralateral hemispheres in a brain slice from an Alb-mSc-ST mouse expressing astrocytic ECS-dSC (left). Scale bar, 100 μm. Quantification of the fluorescence intensity in the two hemispheres (right). Unpaired t-test: ****p < 0.0001 (n = 3 mice).

(f) Entropy levels of cortex and striatum in brain slices from ECS-dSC-expressing Alb-mSc-ST mice injected with the BBB-leakage inducer mannitol or saline. Mann Whitney test: **p = 0.0022 (n = 3 mice, 4 regions each).

All graphs show means ± SEM, apart for violin plots in (d, right).