Clostridium perfringens epsilon toxin induces blood brain barrier permeability via caveolae-dependent transcytosis and requires expression of MAL

Abstract

Clostridium perfringens epsilon toxin (ETX) is responsible for causing the economically devastating disease, enterotoxaemia, in livestock. It is well accepted that ETX causes blood brain barrier (BBB) permeability, however the mechanisms involved in this process are not well understood. Using in vivo and in vitro methods, we determined that ETX causes BBB permeability in mice by increasing caveolae-dependent transcytosis in brain endothelial cells. When mice are intravenously injected with ETX, robust ETX binding is observed in the microvasculature of the central nervous system (CNS) with limited to no binding observed in the vasculature of peripheral organs, indicating that ETX specifically targets CNS endothelial cells. ETX binding to CNS microvasculature is dependent on MAL expression, as ETX binding to CNS microvasculature of MAL-deficient mice was not detected. ETX treatment also induces extravasation of molecular tracers including 376Da fluorescein salt, 60kDA serum albumin, 70kDa dextran, and 155kDA IgG. Importantly, ETX-induced BBB permeability requires expression of both MAL and caveolin-1, as mice deficient in MAL or caveolin-1 did not exhibit ETX-induced BBB permeability. Examination of primary murine brain endothelial cells revealed an increase in caveolae in ETX-treated cells, resulting in dynamin and lipid raft-dependent vacuolation without cell death. ETX-treatment also results in a rapid loss of EEA1 positive early endosomes and accumulation of large, RAB7-positive late endosomes and multivesicular bodies. Based on these results, we hypothesize that ETX binds to MAL on the apical surface of brain endothelial cells, causing recruitment of caveolin-1, triggering caveolae formation and internalization. Internalized caveolae fuse with early endosomes which traffic to late endosomes and multivesicular bodies. We believe that these multivesicular bodies fuse basally, releasing their contents into the brain parenchyma.

A Norrin/Wnt surrogate antibody stimulates endothelial cell barrier function and rescues retinopathy

“…Among the genes regulated by both NDP and F4L5.13, Angpt2 was previously shown to be induced by NDP treatment of cultured human retinal microvascular endothelial cells and to mediate, at least in part, the NDP-mediated increase in proliferation (Ohlmann et al, 2010). Another gene strongly induced by both NDP and F4L5.13 was the membrane protein MAL that was previously identified as the receptor for the Clostridium perfringens epsilon toxin (ETX) on endothelial cells and shown to be required for the ETX effects on blood-brain barrier permeability (Linden et al, 2019).…”

The MAL Protein, an Integral Component of Specialized Membranes, in Normal Cells and Cancer

“…ETX binding is dependent on MAL expression, since no binding to the CNS microvasculature of MAL KO mice was detected. It has been proposed that the effect of ETX on blood–brain barrier permeability in mice involves caveolae-dependent internalization induced by the binding of ETX to MAL at the vessel lumen membrane, and that this triggers the fusion of internalized caveolae with endosomes, the formation of MVBs, their transport across the endothelial cell and, finally, the possible release of exosomes into the opposite membrane, at the brain parenchyma [ 159 ].…”

Echovirus-30 Infection Alters Host Proteins in Lipid Rafts at the Cerebrospinal Fluid Barrier In Vitro

“…In one study, clathrin-mediated endocytosis was required for E-7 to enter polarized Caco-2 cells, and therefore clathrin and dynamin inhibition prevented the entry of the virion in the cells [ 55 ]. Another paper revealed that increased brain barrier permeability was induced by Clostridium perfringens toxin via caveolae dependent transcytosis [ 56 ]. Still, our results showed that clathrin/dynamin blocking had no influence on the damage caused by E-30 on HIBCPP cells.…”