Fluorescence Study of Lipid-based DNA Carriers Properties: Influence of Cationic Lipid Chemical Structure

We report here a study on the physicochemical properties of cationic phospholipids liposomes used for lipoplex formulation and DNA transfer. The original cationic phospholipids synthesized in our laboratory are first presented with the liposome formulation process. The second part deals with the liposomes fusogenic properties studied by fluorescence resonant energy transfer (FRET). The nature of the cationic polar head and the formulation with or without a neutral colipid have a great influence on the FRET signal. The third part reports the study of the viscosity of the liposome by fluorescence anisotropy measurements. It has been observed that the vectors having a saturated lipid chain exhibit a more pronounced anisotropy than those having unsaturated lipid chains. Finally, liposomes formed by a mixture of phospholipids and DC-Chol (a rigid lipid) leads to increase the anisotropy denoting a more rigid liposome.     

番茄红素脂质体的制备工艺

采用逆相蒸发法和薄膜法两种方法制备番茄红素脂质体.筛选出制备高包封率脂质体的工艺为:卵磷脂与胆固醇之比为3:1,温度为35℃,番茄红素与磷脂比例为1:6,旋转速度为200r/min,此条件下番茄红素脂质体的最高包封率可达71.26%.     

Review of binding methods and detection of Al(III) binding events in trypsin and DL-DPPC liposomes by a general thermodynamic model

In this paper are reviewed some of the most useful binding formalisms which have been developed in order to investigate the diversity of the commonly encountered receptor-ligand systems. Particularly, our attention is focused on the presentation and application of a general and rigorous thermodynamic treatment which explains in a simple and coherent way the non-linear profiles observed in the experimental binding, Scatchard, Hill and Adair plots. By applying this model, both qualitative and detailed quantitative investigations were carried out on the binding process of Al(III) to trypsin and dl-dipalmitoylphosphatidylcholine (DPPC) liposomes. Particularly, it has been demonstrated that: Al(III) interacts with liposomes in two binding sites with very different dissociation constants. The first one has been calculated to be 0.0168 μmol l⁻¹ and the second 2.833 μmol l⁻¹. The first is referred to the preferential interaction of Al³⁺ with the polar head of the phospholipid, while the second most likely regards the interaction of Al³⁺ with other peripheral sites. On the other hand, Al³⁺ interacts with trypsin in two types of binding sites. The first binding site with K_1,1=0.2531 μmol l⁻¹ and the second with K_2,1=1.424 μmol l⁻¹.     

Interaction of Na2B12H11SH with liposomes: Influence on zeta potential and particle size

The interaction of Na₂B₁₂H₁₁SH (BSH) with liposomes has been studied. BSH is a compound used clinically in boron neutron capture therapy of glioblastoma, and is known to enter tumor cells. Liposomes were used as a model for studying the interaction of BSH with cell membranes. BSH led to changes in the zeta potential of liposomes consisting of DODAB (dioctadecyldimethylammonium bromide) alone or with DOPC (dioleylphosphatidylcholine) or DOPE (dioleylphosphatidylethanolamine). It also led to changes of the size of DODAB liposomes, with a maximum size at small zeta potentials. A firm binding of BSH with the head groups of the lipid must be assumed.     

Solubilization of egg phosphatidylcholine liposomes by sodium taurocholate: partition behavior and morphology

Solubilization process of egg yolk phosphatidylcholine (EggPC) liposomes by sodium taurocholate (TC) was performed using membrane dialysis technique. Turbidity, apparent particle size, Cl⁻ permeability, membrane fluidity measured by electron spin resonance spectroscopy (ESR), and structural changes observed by freeze-fracture electron microscopy were investigated. The concentrations of TC were analyzed by high-performance liquid chromatography to determine the incorporation of TC in membrane phase and apparent partition coefficient. When TC concentration in water phase (D _w) was lower than 1.41 mM, or effective ratio of TC to EggPC (R _e) was less then 0.14, the partition coefficient was independent of the TC concentration and was 90 M⁻¹, where the membrane permeability of Cl⁻ was rather small. Upon increasing the concentrations in the range of 1.41 mM≤D _w<3.10 mM (0.14≤R _e<0.21), the apparent partition coefficient decreased and membrane permeability markedly increased. The coexistence of small vesicles and bilayer fragments were observed in this region. At 3.10 mM≤D _w<3.53 mM (0.21≤R _e<0.59), the steep decrease in turbidity was accompanied with the gradual increase in apparent partition coefficient, suggesting the formation of mixed micelles. The freeze-fracture electron micrographs confirmed the transformation of saturated bilayers to wormlike mixed micelles. When a further increase in TC (D _w≥3.53 or R _e≥0.59) was applied, the turbidity and apparent particle sizes became small. Transformation of the wormlike structure to ellipsoid mixed micelles was observed. The ESR spectra showed that TC/EggPC mixed micelles still retain a somewhat orderly structure.     

Adiabatic compressibility of red blood cell membrane: influence of skeleton

Measurements of ultrasound velocity and density were used for determination of the adiabatic compressibility of red blood cells (RBC) during detachment of the membrane skeleton. Skeleton detachment was induced by addition of nystatin into a low ionic strength RBC suspension resulting in an increase (10%) of the ultrasound velocity concentration increment, [u], while the specific volume of cells, phi(v) did not change significantly. Changes of the concentration increment had rather long kinetics and were not completed even after 60 min. Both [u] and phiV values were used for calculation of the specific apparent adiabatic compressibility of RBC, phiK/beta0. The value of the specific apparent compressibility decreases following addition of nystatin. This corresponds to an increase in the volume elastic modulus of RBC membranes during detachment of the membrane skeleton. Control experiments with large unilamellar liposomes at conditions similar to that performed with the RBC did not reveal significant changes of [u] after the addition of nystatin. Our results show that the role of the membrane skeleton probably consists in maintaining higher compressibility of the RBC membranes. This may partly provide conditions for conformational changes of RBC membrane proteins.     

Effect of variation in the chain length and number in modulating the interaction of an immunogenic lipopeptide with biomembrane models

A differential scanning calorimetry study was carried out to investigate the effect exerted by immunogenic synthetic lipopeptides obtained by the conjugation of LCMV_33–41 peptide with lipoamino acids (Laas) bearing different alkyl chain lengths (C₁₂ and C₁₆) and number of chains (2 × C₁₂) on the thermotropic behaviour of dimyristoylphosphatidylcholine (DMPC) liposomes. The aim of this work was to study the ability of these compounds to be carried by a liposomal system and released to a biomembrane model.

The examined compounds caused variations of the thermotropic parameters that characterise the liposomal system (transition temperature, T_m and enthalpy variation, ΔH), and interacted with the biomembrane models in different way. The interaction was found to be modulated by the length and number of chains present in the examined compounds. In fact, the compounds with higher number of lipid chain showed a stronger interaction with the biomembrane models with respect to the pure peptide and the compounds with a single lipid chain. These results suggest that the lipoamino acid moiety could favour the peptide to be carried by the liposomal system and released to biomembrane.     

Tripolyphosphate-sensitive egg phosphatidylcholine liposomes incorporating hydrophobically modified poly(ethylene imine)

Liposomes, which release their contents in answer to tripolyphosphate (TPP, a penta-anion), were prepared by immobilizing hydrophobically modified poly(ethylene imine) (HmPEI) on the surface of egg phosphatidylcholine (egg PC) liposome. HmPEI was prepared by covalently attaching decanoyl chloride to PEI through a condensation reaction. According to the ¹H NMR spectrum, the number of decanoyl chloride per one molecule of PEI was about 21, and HmPEI was air/water interface-active. HmPEI could readily complex with TPP in HEPES buffer (30 mM, pH 7.0), confirmed by Fourier transformed infrared spectrophotometer spectroscopy. The complexation increased with increasing the concentration of HmPEI and TPP, investigated through the measurement of optical density and light scattering intensity. Liposomes incorporating HmPEI were prepared by a film hydration and sonication method. The liposomes were multi-lamellar vesicles, observed on transmission electron microscope. Liposomes free of HmPEI did not release calcien when they were mixed with TPP. Liposomes whose egg PC/HmPEI was relatively low (e.g., 20:1 and 20:2) released calcein but not extensively (less than 10%) when mixed with TPP. Liposomes whose egg PC/HmPEI was relatively high (e.g., 20:4 and 20:20) released calcein extensively. For example, when the liposomes with lager amount of HmPEI were mixed with TPP so that HmPEI/TPP weight ratio was 8:1, the release degree in 60 sec was more than 70%. HmPEI can complex with TPP through electrostatic interaction and the complexation was thought to cause perturbation in the liposomal membranes and trigger the release.     

Dynamics of liposomes in the fluid phase

We review the currently available material on the morphology and dynamics of phospholipids assembled into liposomes. Key information obtained from neutron scattering, nuclear magnetic resonance (NMR), and other techniques plays a crucial role in understanding the vital role of lipids in sustaining life in living organisms. We concentrate on the dynamics in the biologically important fluid phase in the time range from picoseconds to seconds, which includes a discussion of the center of mass diffusion of liposomes, membrane fluctuations; and lateral, rotational, and flip-flop motions of the lipids. We emphasize on the sensitivity of the dynamics on interactions with a variety of biologically relevant molecules such as cholesterol. By a comparison of data from literature, we witness a good agreement of the results from different techniques and studies.     

Influence of cyclodextrins on the physical stability of DPPC-liposomes

This paper reports on the physical stability of DPPC-(dipalmitoyl phosphatidyl choline) liposomes in various aqueous dispersions and its control by uncharged polymers. The effect of natural (-, β-, γ-) cyclodextrins (CDs) on the stability of bare and polymer-bearing liposomes and also, the attachment of the CD molecules and the macromolecules, respectively, to the DPPC-bilayers of small unilamellar vesicles (SUV) were studied.

It was found that above a CD/DPPC ratio, each cyclodextrin caused a definite destruction in the phospholipid bilayers. The extent of membrane destabilization due to a cyclodextrin closely related to the amount of the CD molecules bound to the DPPC-bilayers.

The polymer-coated liposomes formulated by incorporating a dissolved homopolymer or copolymer into the phospholipid bilayer of the vesicles exhibited higher physical stability. Uncharged polymers effectively hindered the disintegration of the liposomal membranes brought about by the CD molecules. The polymer layers formed around the phospholipid bilayers ensured an enhanced steric stabilization for the DPPC-liposomes. Methylcellulose (MC) with high molecular mass and a polyvinyl alcohol-co-vinyl propional copolymer alike exhibited efficient stabilizing effect.