Membrane interactions of ternary phospholipid/cholesterol bilayers and encapsulation efficiencies of a RIP II protein

• Published in: Colloids and surfaces, B, Biointerfaces B, Biointerfaces, 2008-07, Vol.64 (2), p.284-296
• Main Authors: Manojlovic, V., Winkler, K., Bunjes, V., Neub, A., Schubert, R., Bugarski, B., Leneweit, G.
• Format: Article
• Language: English
• Subjects: 1,2-Dipalmitoylphosphatidylcholine - chemistry; Cellular Structures - chemistry; Cellular Structures - metabolism; Cholesterol; Cholesterol - chemistry; Cryoelectron Microscopy; DOPC; DPPC; Drug Stability; Enzyme-Linked Immunosorbent Assay; Extrusion; Freezing; Lipid Bilayers - chemistry; Liposomes; Liposomes - chemistry; Liposomes - isolation & purification; Liposomes - metabolism; Lysis tension; Mistletoe lectin; Particle Size; Phosphatidylcholines - chemistry; Phospholipids - chemistry; Pressure; Receptor-Interacting Protein Serine-Threonine Kinase 2 - chemistry; Receptor-Interacting Protein Serine-Threonine Kinase 2 - metabolism; Receptor-Interacting Protein Serine-Threonine Kinase 2 - ultrastructure; Santalales; Time Factors
• ISSN: 0927-7765; 1873-4367
• DOI: 10.1016/j.colsurfb.2008.02.001

Membrane interactions of liposomes of ternary phospholipid/cholesterol bilayers are investigated. These interactions lead to discoidal deformations and regular aggregations and are strongly enhanced by the presence of mistletoe lectin (ML), a RIP II type protein. The encapsulation of ML into liposomal nanocapsules is studied with a systematic variation of the lipid composition to monitor its effect on the physical properties: entrapment, mean size, morphology, and stability. Extrusion of multilamellar vesicles through filters 80 nm pore size was used for the generation of liposomes. The mean sizes of liposomes ranged between 120 and 200 nm in diameter with narrow size distributions. The increase in flow rate with pressure for three dioleoylphosphatidylcholine (DOPC)/cholesterol (Chol)/dipalmitoylphosphatidylcholine (DPPC) lipid mixtures was linear and allowed to extrapolate to the minimum burst pressure of the liposomal bilayers. From the minimum pressures P min, the bilayer lysis tensions γ l were determined. The increase in P min and γ l with an increasing content of a saturated phosopholipid (DPPC) indicates that DPPC increases the mechanical strength of lipid bilayers. Apparently, DPPC, like cholesterol, leads to a less compressible surface and a more cohesive membrane. After preparation, vesicle solutions were purified by gel permeation chromatography to separate encapsulated ML from free ML in the extravesicular solution. Purified liposomes were then characterized. The content of entrapped and adsorbed ML was measured using ELISA. Repetitive freezing/thawing cycles prior to extrusion significantly increased ML uptake. On the contrary, adsorption was not affected neither by lipid composition, nor concentration and preparation. Differences in experimental encapsulation efficiency only reflect the differences in the mean vesicle sizes of the different samples as is revealed by a comparison to a theoretical estimate. Cryo-transmission electron microscopy (Cryo-TEM) images show that beside spherical, single-walled liposomes, there is a considerable fraction of discoidally deformed vesicles. Based on our results and those found in the literature, we speculate that the flattening of the vesicles is a consequence of lipid phase separation and the formation of condensed complexes and areas of different bending elasticities. This phenomenon eventually leads to agglomeration of deformed liposomal structures, becoming more pronounced with the increase in the rel