Linden, Jennifer R. Flores, Claudia Schmidt, Eric F. Uzal, Francisco A. Michel, Adam O. Valenzuela, Marissa Dobrow, Sebastian Vartanian, Timothy CAV1 expression is necessary for ETX induced BBB permeability but not ETX binding to brain microvasculature. <p>(A) To determine if CAV1 was necessary for ETX binding to brain microvasculature, cerebellum cryosections from <i>Cav1+/+</i> and <i>Cav1-/-</i> were probed with epsilon protoxin (proETX). ProETX binding was detected by and anti-ETX antibody (red). FITC-BSL1 was used to identify vasculature (green). Note that proETX is observed binding to the microvasculature in the molecular layer (ML) as well as myelin in the granular layer (GL) in both <i>Cav1</i>+/+ and <i>Cav1</i>-/- mice. (B) Higher magnification images of areas in white boxes in image A. (C) Quantification of proETX binding to vasculature in cerebellum gray matter in <i>Cav1</i>+/+ and <i>Cav1</i>-/- mice. proETX fluorescence was normalized to BSL1 fluorescence. Results expressed as Mean ± STDEV, *p value determined by T-Test, n = 4–7. D) To evaluate CAV1’s role in ETX induced BBB permeability, <i>Cav1</i>+/+ <i>Cav1</i>-/- were treated with or without 5ng of ETX per gram of body weight for up to 180 minutes. Mice were perfused with PBS, brains harvested, and cryosectioned. BBB permeability was accessed by immunohistochemistry staining for endogenous mouse IgG in the pericollosal white matter of sagittal sections. Results are normalized to individual genotype controls and expressed as IgG Extravasation (% CT). Results expressed as Mean ± STDEV, * p < 0.05 determined by ANOVA, n = 2–3 mice per group. (E) Representative images from ETX treated <i>Cav1</i>+/+ and <i>Cav1</i>-/- treated mice. Note in ETX treated <i>Cav1</i>+/+ mice, endogenous IgG (red) can be observed leaking for large medullary veins (white arrows) and smaller capillaries (white arrow heads). FITC-BSL1 (green) was used to identify vasculature.</p> RAB;MAL Clostridium perfringens epsilon toxin;Clostridium perfringens epsilon toxin;endosome;EEA;Internalized caveolae fuse;lipid raft-dependent vacuolation;60 kDA serum albumin;155 kDA IgG;multivesicular bodies fuse basally;CNS microvasculature;caveolae-dependent transcytosis;ETX causes blood brain barrier;70 kDa dextran;ETX causes BBB permeability;376 Da fluorescein salt;ETX binding;multivesicular bodies;blood brain barrier permeability;ETX-induced BBB permeability;exhibit ETX-induced BBB permeability 2019-11-08
    https://plos.figshare.com/articles/figure/CAV1_expression_is_necessary_for_ETX_induced_BBB_permeability_but_not_ETX_binding_to_brain_microvasculature_/10277531
10.1371/journal.ppat.1008014.g006