Uptake of apolipoprotein E fragment coupled liposomes by cultured brain microvessel endothelial cells and intact brain capillaries

The suitability of surface modified liposomes as drug carriers for brain-specific targeting was investigated using apolipoprotein E fragments as brain-directed vectors. Liposomes coated with polyethylene glycol-2000 (sterically stabilized, PEGylated liposomes) were prepared from hydrogenated egg phosphatidylcholine, cholesterol, and a PEG-derivatized phospholipid. Liposomes were covalently coupled to a peptide of 26 amino acids length, derived from the binding site of human apolipoprotein E4 (ApoE4) and a peptide of random amino acid sequence, respectively. Rhodamine-labeled dipalmitoylphosphatidylethanolamine was incorporated into the lipid bilayer in order to visualize the liposomal interaction with brain capillary endothelial cell monolayers. The interaction of the liposomes with monolayers of porcine brain capillary endothelial cells (BCEC), the rodent cell line RBE4, and freshly isolated porcine brain capillaries was studied by means of confocal laser scanning fluorescence microscopy. In contrast to random peptide coupled liposomes, the ApoE4-fragment coupled liposomes were rapidly taken up by cultured BCECs and RBE4 cells. Uptake could be inhibited by ApoE4, free peptide, and antibodies against the LDL receptor in a concentration-dependent manner. The results indicate that the liposomes are internalized via the LDL receptor, which is expressed at the blood-brain barrier. In conclusion, liposomes coupled to ApoE4 fragments are taken up into brain endothelium via an endocytotic pathway and may therefore be a suitable carrier for drug delivery to the brain.