SPECIFIC BINDING OF A MUTATED FRAGMENT OF CLOSTRIDIUM PERFRINGENS ENTEROTOXIN TO ENDOTHELIAL CLAUDIN-5 AND ITS MODULATION OF CEREBRAL VASCULAR PERMEABILITY

The vertebrate blood-brain barrier (BBB) creates an obstacle for central nervous system-related drug delivery. Claudin-5 (Cldn5), expressed in large quantities in BBB, plays a vital role in restricting BBB permeability. The C-terminal domain of Clostridium perfringens enterotoxin (cCPE) has been verified as binding to a subset of claudins (Cldns). The Cldn5-binding cCPE(194-319) variant cCPE(Y306W/S313H) was applied in this study to investigate its ability to modulate the permeability of zebrafish larval BBB. In vitro results showed that cCPE(Y306W/S313H) is able to bind specifically to Cldn5 in murine brain vascular endothelial (bEnd. 3) cells, and is transported along with Cldn5 from the cell membrane to the cytoplasm, which in turn results in a reduction in transendothelial electrical resistance (TEER). Conversely, this effect can be reversed by removal of cCPE(Y306W/S313H). In an in vivo experiment, this study estimates the capability of cCPE(Y306W/S313H) to modulate Cldn5 using a rhodamine B-Dextran dye diffusion assay in zebrafish larval BBB. The results show that cCPE(Y306W/S313H) co-localized with Cldn5 in zebrafish cerebral vascular cells and modulated BBB permeability, resulting in dye leakage. Taken together, this study suggests that cCPE(Y306W/S313H) has the capability - both in vitro and in vivo - to modulate BBB permeability temporarily by specific binding to Cldn5. (C) 2016 IBRO. Published by Elsevier Ltd. All rights reserved.