UPTAKE OF LIPOSOMES WHICH INCORPORATE A GLYCOPEPTIDE FRACTION OF ASIALOFETUIN BY HepG(2) CELLS

We examined the interaction between liposomes which incorporate a fraction triantennary glycopeptide (AF(2)) of asialofetuin and human hepatoma cells (HepG(2)) in vitro. HepG(2) cells are known to express the asialoglycoprotein receptor. For liposome preparation AF(2) was cleaved from asialofetuin, purified and conjugated with different length (C(12),C(16) and C(18)) fatty acids (FA). The conjugates were subsequently incorporated into pre-formed sonicated liposomes using a mild cholate incubation method. Interactions between AF(2)/FA-liposomes as well as control-liposomes (with no ligand) and cells (in the presence of serum) were measured at different lipid doses after incubating HepG(2) cells with liposomes at 4 degrees C and 37 degrees C, in the absence and presence of galactose, and also evaluated by fluorescence microscopy. More extensive studies were performed with the AF(2)/C(18)-liposomes which were previously found to incorporate higher amounts of ligand and be the most stable of the formulations prepared. Results from both, morphological and quantitative studies, demonstrate that AF(2)/C(16) and especially AF(2)/C(18)-liposomes are bound and taken up by the cells by a galactose specific mechanism. The AF(2)/C(12)-liposomes-which were previously found to incorporate low amounts of ligand in a non-stable way- were taken up by the cells in amounts similar to those of the control liposomes (without ligand) while this uptake was not reduced by galactose and therefore possibly non-specific. The intracellular localization of AF(2)/C(18)-liposomes was further evidenced by intracellular acidification using NH(4)Cl. These conclusions, justify the importance of further in vivo studies in order to demonstrate the capability of the proposed system to target hepatocytes.