Patchy cationic liposomes as fusogenic particles.

Abstract

Membrane fusion facilitates transport of molecules and particles across membranes and is thus the key to many biological processes such as viral infection. Membrane fusion has also been utilized for intracellular delivery of macromolecules using, for instance, fusogenic liposomes. These liposomes that often contain substantial amounts of cationic lipid DOTAP can, however, lead to toxicity. Recently, membrane phase-separation was explored to enhance fusion of cationic liposomes. This study aims to further explore the use of phase-separation to create patchy cationic liposomes with high fusogenicity and low toxicity. We apply small cationic liposomes, of homogenous and phase-separating compositions, and assess their fusogenicity using model membranes and cell membranes. Giant liposomes of PC or PC/PS were used as target model membranes where their fusion with cationic liposomes was evaluated using fluorescence microscopy. Two cell lines of U87 and HEK293 were selected due to their distinct surface charges and their membrane fusion was assessed by flow cytometry. For patchy cationic liposomes, we applied ternary composition DOPC:DPPC:Chol with three different molar ratios for incorporation of DOTAP. Keeping DOTAP mol% constant while changing the phase-separating membrane composition allowed us to examine the effect of DOTAP surface density on liposomal fusogenicity. When using homogeneous cationic liposomes, our results in both model and cellular membrane studies suggested that while increasing DOTAP content in these liposomes could increase their fusogenicity, the maximal fusion efficacy was also affected by the surface charge of the target membrane. Phase-separating liposomes showed higher fusion compared to their homogenous counterparts with similar DOTAP content. Lastly, among phase-separating compositions, the one that led to highest local concentration of DOTAP on liposomes showed the highest level of fusion into target model membranes. These findings show the potential of patchy cationic liposomes as highly fusogenic particles for intracellular delivery.