STUDY OF SULFATIDE-CONTAINING LIPOSOMES FOR DELIVERY TO CELLS

Abstract

Phospholipid liposomes have been studied extensively as vehicles for targeting delivery to tissues and/or cells. Several investigations suggest that a glycosphingolipid, sulfatide, can be an active component in liposome-mediated delivery of macromolecules into the brain. The thesis ventures to design and characterize the sulfatide-containing 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) liposomes. One of our objectives is to obtain the stable liposomes with increased half-life in the blood circulation. On the other hand, if endocytosis is the major pathway for cells to take up liposomes, the stability of these liposomes should be pH-sensitive in order to escape the lysosomal degradation. It has been found that increasing sulfatide concentration up to 30 mol% enhances the stability of the sulfatide-containing DOPE liposomes. This effect of sulfatide is leveled off at higher concentrations. The pH sensitivity of the liposomes is retained at 30 mol% sulfatide in the presence of human plasma. It has been shown that the sulfatide-induced increase in bilayer surface hydration and decrease in interchain hydration play an essential role in stabilizing the DOPE/sulfatide bilayer vesicles, while the importance of the condensing effect of the glycosphingolipid is less significant. Furthermore, the pH-sensitivity of these liposomes is associated with the partial dehydration at the bilayer surface exposed to acidic media. Under this condition, the DOPE molecules may have a tendency to take a non-bilayer configuration that would destabilize the structure of the bilayer vesicles. Again, no further change in headgroup hydration and interchain hydration near the bilayer center is observed at sulfatide concentrations higher than 30 mol%. The correlation between the concentration-dependent effects of sulfatide on membrane hydration, stability and pH-sensitivity are explained by taking into account the electrostatic and geometrical properties of the sulfated galactosyl headgroup of sulfatide. For cytoplasmic delivery of liposomal contents, it is important for the liposomes to be able to fuse with the endosomal membranes. It has been found that the fusogenic capability of DOPE/sulfatide liposomes is reduced at increasing concentration of the glycosphingolipid. This inhibitory effect is more obvious at sulfatide concentrations higher than 30 mol%, where the fusion is no longer pH-sensitive. Similar inhibitory effect is observed in other glycosphingolipid-containing liposomes. The results are consistent with an increasing steric hindrance to membrane fusion at higher sulfatide concentrations. Interestingly, the pH sensitivity of the sulfatide-containing liposomes is retained when they are allowed to fuse with biological membranes (synaptosomes) in the absence of ca2+, probably by a mechanism involving protein mediation. The cellular uptake of DOPE/sulfatide liposomes has been investigated by incubating the liposomes with various cell lines. It has been shown that these liposomes are specifically recognizable by CCF cells originated from a human astrocytoma. Presence of unsaturated PE molecules is necessary for efficient cellular uptake. All evidence obtained indicates that endocytosis, rather than direct fusion between the liposomes and the plasma membrane of the target cells, is the major pathway of cellular uptake of DOPE/sulfatide liposomes. A report gene has been condensed in the sulfatide-containing liposomes and delivered successfully to the CCF cells for expression. Taken together, the pH-sensitive DOPE/sulfatide liposomes are promising carriers for delivery of drugs/genes into cells, which can be developed further for various clinical applications.