N-isopropylacrylamide based nanogels crossing the blood-brain barrier: evidence of high in vitro internalization in human endothelial cells and in vivo permeation in zebrafish

The development of effective neurological treatments remains a challenge due to the low permeation across the blood-brain barrier (BBB). The use of nanoparticles for brain drug delivery is promising, but the level of BBB crossing remains low, typically around 0.5-3%. We report the synthesis of negatively charged N-isopropylacrylamide nanogels (diameter 10 nm) displaying in vitro crossing in a human BBB model nearly 5-fold higher than previously reported, and permeating the brain parenchyma of healthy zebrafish with a fully formed BBB. The methylenebisacrylamide crosslinked nanogels, incorporating N-acryloyl-L-proline as pH-responsive unit (electrophoretic mobility = − 0.24 ± 0.05 μm·cm·V-1·s-1), were fluorescently labelled with (4-(2-acryloylaminoethylamino)-7-nitro-2,1,3-benzoxadiazole, to allow in vitro and in vivo tracking. The nanoparticles did not disrupt the in vitro human BBB model up to 0.1 mg/mL, obtained from CD34 positive-derived endothelial cells in co-culture with pericytes, displaying 13.4% permeation at 24 hours; internalization in the BBB endothelial cells was confirmed by confocal microscopy, via clathrin-mediated endocytosis. The ability of nanogels to cross the BBB in vivo without inducing a toxic effect was confirmed in zebrafish, up to 14 days post fertilization.