Vesicle Suspension Research Articles

Phase separation and collapse in almost density matched depletion induced colloidal gels in presence and absence of air bubbles: An MRI imaging study

HypothesisVesicle-polymer dispersions are found in drug-delivery systems and consumer products but undergo phase separation. Previous studies of phase separation have focussed on systems with high density differences between continuous and vesicular phases. In this study, we investigate phase separation in vesicle-polymer mixtures with very small density differences, in the presence and absence of air bubbles. ExperimentsMagnetic resonance (MR) imaging, X-ray Computed Tomography and rheological measurements are reported which characterise the properties and stability of vesicle suspensions composed of the cationic surfactant, diethylesterdimethyl ammonium chloride, mixed with non-adsorbing polymer. 1H T2 MR relaxation images are employed to observe phase separation, for a range of vesicle-polymer mixtures, which are analysed using Moran’s I spatial autocorrelation to quantify the extent and rate of phase separation. FindingsIt was found that in presence of air bubbles, phase separation follows a compression/collapse mechanism, typical of colloidal gels with large density differences between the phases. Without air bubbles, phase separation develops through the formation of tiny cracks and fractures in the samples. MRI enabled visualisation of the evolution of phase separation inside highly turbid samples. The rate of phase separation was found to generally increase with increasing polymer concentration and decrease with increasing vesicle volume fraction.

Validation of conductivity tensor imaging using giant vesicle suspensions with different ion mobilities

BackgroundElectrical conductivity of a biological tissue at low frequencies can be approximately expressed as a tensor. Noting that cross-sectional imaging of a low-frequency conductivity tensor distribution inside the human body has wide clinical applications of many bioelectromagnetic phenomena, a new conductivity tensor imaging (CTI) technique has been lately developed using an MRI scanner. Since the technique is based on a few assumptions between mobility and diffusivity of ions and water molecules, experimental validations are needed before applying it to clinical studies.MethodsWe designed two conductivity phantoms each with three compartments. The compartments were filled with electrolytes and/or giant vesicle suspensions. The giant vesicles were cell-like materials with thin insulating membranes. We controlled viscosity of the electrolytes and the giant vesicle suspensions to change ion mobility and therefore conductivity values. The conductivity values of the electrolytes and giant vesicle suspensions were measured using an impedance analyzer before CTI experiments. A 9.4-T research MRI scanner was used to reconstruct conductivity tensor images of the phantoms.ResultsThe CTI technique successfully reconstructed conductivity tensor images of the phantoms with a voxel size of 0.5times 0.5times 0.5hbox { mm}^3. The relative L^2 errors between the conductivity values measured by the impedance analyzer and those reconstructed by the MRI scanner was between 1.1 and 11.5.ConclusionsThe accuracy of the new CTI technique was estimated to be high enough for most clinical applications. Future studies of animal models and human subjects should be pursued to show the clinical efficacy of the CTI technique.

Shape transition and hydrodynamics of vesicles in tube flow

The steady motion and deformation of a lipid-bilayer vesicle translating through a circular tube in low Reynolds number pressure-driven flow are investigated numerically using an axisymmetric boundary element method. This fluid-structure interaction problem is determined by three dimensionless parameters: reduced volume (a measure of the vesicle asphericity), geometric confinement (the ratio of the vesicle effective radius to the tube radius), and capillary number (the ratio of viscous to bending forces). The physical constraints of a vesicle -- fixed surface area and enclosed volume when it is confined in a tube -- determine critical confinement beyond which it cannot pass through without rupturing its membrane. The simulated results are presented in a wide range of reduced volumes [0.6, 0.98] for different degrees of confinement; the reduced volume of 0.6 mimics red blood cells. We draw a phase diagram of vesicle shapes and propose a shape transition line separating the parachute-like shape region from the bullet-like one in the reduced volume versus confinement phase space. We show that the shape transition marks a change in the behavior of vesicle mobility, especially for highly deflated vesicles. Most importantly, high-resolution simulations make it possible for us to examine the hydrodynamic interaction between the wall boundary and the vesicle surface at conditions of very high confinement, thus providing the limiting behavior of several quantities of interest, such as the thickness of lubrication film, vesicle mobility and its length, and the extra pressure drop due to the presence of the vesicle. This extra pressure drop holds implications for the rheology of dilute vesicle suspensions. Furthermore, we present various correlations and discuss a number of practical applications.

Low‐wavenumber Raman scattering of phospholipid bilayers in fluid, ripple, and gel phases

AbstractLow‐wavenumber Raman spectra of the aqueous suspension of multilamellar vesicles made of 1,2‐dimyristoyl‐sn‐glycero‐3‐phosphocholine have been studied. The Raman susceptibility of phospholipid bilayers was experimentally found for the gel, ripple, and fluid lipid phases. Comparison of the Raman susceptibility with the inelastic neutron and X‐ray scattering data evidences that the Raman spectrum reflects the in‐plane phonon density of states. Also, the Raman susceptibility reveals peaks below ~ 30 cm−1 associating with the off‐plane modes. These peaks were found to be sensitive to the lipid phase. In particular, the ripple spectrum included the off‐plane modes of the fluid and gel phases. The low‐wavenumber Raman spectroscopy becomes a powerful method for studying THz vibrational modes of phospholipid bilayers, corresponding to nanometer response of the bilayers.

Rheology of a confined vesicle suspension

The effect of an off-center stable steady state of a vesicle (a model of red blood cells) in a bounded shear flow is investigated numerically. It is found that this new solution (off-center) strongly alters the rheological properties of dilute as well as concentrated vesicle suspensions.

Fast and slow biomembrane solubilizing detergents: Insights into their mechanism of action

Detergents are water-soluble amphiphiles. Above a critical concentration they self-organize in micelles and in the presence of phospholipids mixed micelles are formed. Much information is available on the structure of these self-assemblies and on the thermodynamics of their formation. The aim of this study was to deepen our understanding of the mechanisms of solubilization. Solubilization of lipid vesicles made of egg phosphatidylcholine (PC) by twenty one commercially available, structurally heterogeneous detergents, has been assessed by a decrease in turbidity of the vesicle suspension. Both steady-state and time-resolved measurements have been performed. The results show that the detergents under study fall into one of two categories, namely fast-solubilizing and slow-solubilizing detergents. This categorization is independent of detergent concentration, i.e. a “slow” cannot be converted into a “fast” surfactant by increasing its bulk concentration. 31P-NMR spectra indicate that slow-acting detergents cause either a gradual, monotonic micellization of bilayers (sodium dodecyl sulphate), or formation of more complex, perhaps non-lamellar, non-micellar intermediates (dodecylmaltoside). In contrast, fast detergents (e.g. Triton X-100) cause lysis and reassembly of vesicles before bulk solubilization takes place. These results support the idea that membrane solubilization by detergents is rapid only when surfactant transbilayer (flipping) motion is easy.

Machine learning acceleration of simulations of Stokesian suspensions.

Particulate Stokesian flows describe the hydrodynamics of rigid or deformable particles in Stokes flows. Due to highly nonlinear fluid-structure interaction dynamics, moving interfaces, and multiple scales, numerical simulations of such flows are challenging and expensive. Here, we propose a generic machine-learning-augmented reduced model for these flows. Our model replaces expensive parts of a numerical scheme with regression functions. Given the physical parameters of the particle, our model generalizes to arbitrary geometries and boundary conditions without the need to retrain the regression functions. It is approximately an order of magnitude faster than a state-of-the-art numerical scheme using the same number of degrees of freedom and can reproduce several features of the flow accurately. We illustrate the performance of our model on integral equation formulation of vesicle suspensions in two dimensions.

Enhancement of optical force acting on vesicles via the binding of gold nanoparticles.

Here we found that gold nanoparticles (AuNPs) enhance the optical force acting on vesicles prepared from phospholipids via hydrophobic and electrostatic interactions. A laser beam was introduced into a cuvette filled with a suspension of vesicles and it accelerated them in its propagation direction via a scattering force. The addition of the AuNPs exponentially increased the velocity of the vesicles as their concentration increased, but polystyrene particles had no significant impact on velocity in the presence of AuNPs. To elucidate the mechanism of the increased velocity, the surface charges in the vesicles and the AuNPs were controlled; the surface charges of the vesicles were varied via the use of anionic, cationic and neutral phospholipids, whereas AuNPs with positive and negative charges were synthesized by coating with citrate ion and 4-dimethylaminopyridine, respectively. All vesicles increased the velocity at different degrees depending on the surface charge. The vesicles were accelerated more efficiently when their charges were opposite those of the AuNPs. These results suggested that hydrophobic and electrostatic interactions between the vesicles and the AuNPs enhanced the optical force. By accounting for the binding constant between the vesicles and the AuNPs, we proposed a model for the relationship between the concentration of the AuNPs and the velocity of the vesicles. Consequently, the increased velocity of the vesicles was attributed to the light scattering that was enhanced when AuNPs were adsorbed onto the vesicles.

Electrohydrodynamics of deflated vesicles: budding, rheology and pairwise interactions

We develop a new boundary integral method for solving the coupled electro- and hydrodynamics of vesicle suspensions in Stokes flow. This relies on a well-conditioned boundary integral equation formulation for the leaky-dielectric model describing the electric response of the vesicles and an efficient numerical solver capable of handling highly deflated vesicles. Our method is applied to explore vesicle electrohydrodynamics in three cases. First, we study the classical prolate–oblate–prolate transition dynamics observed upon application of a uniform DC electric field. We discover that, in contrast to the squaring previously found with nearly spherical vesicles, highly deflated vesicles tend to form buds. Second, we illustrate the capabilities of the method by quantifying the electrorheology of a dilute vesicle suspension. Finally, we investigate the pairwise interactions of vesicles and find three different responses when the key parameters are varied: (i) chain formation, where they self-assemble to form a chain that is aligned along the field direction; (ii) circulatory motion, where they rotate about each other; (iii) oscillatory motion, where they form a chain but oscillate about each other. The last two are unique to vesicles and are not observed in the case of other soft particle suspensions such as drops.

Brillouin study of elastic properties of nanometric phospholipid layers in aqueous suspensions of vesicles

This work is devoted to the development of an experimental technique based on Brillouin spectroscopy for the study of aqueous suspensions of multilayer vesicles of phospholipids at different temperatures using the example of DPPC phospholipid (1,2-dipalmitoyl-sn-glycero-3-phosphocholine). The temperature dependence of the Brillouin line position is studied. A comparison of the behavior of Brillouin lines of polycrystalline ice, dry vesicles, and vesicles suspension reveals the contribution of phospholipid layers to the Brillouin spectrum. Analysis of the data makes it possible to estimate the value of sound velocity of the vesicles membranes and its temperature dependence. [AQ1] [AQ2]

Interactions of valproic acid with lipid membranes of 1,2-dimyristoyl-sn-glycero-3-phosphocholine.

Lipid bilayers of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) were prepared in two forms, as a suspension of multilamellar spherical vesicles and as planar membranes deposited on a conductive solid support. We used Fourier Transformed Infrared (FTIR) and Raman spectroscopic techniques to study the lipid vesicles while the solid supported bilayers were characterized by using electrochemical experiments (cyclic voltammetry and impedance). Valproic acid (Valp) was either present in the solution or incorporated into the lipid structure. As the Valp:DMPC ratio increases the phase transition temperature decreases while the phase transition becomes less marked. Moreover, for the Valp:DMPC complex species a slight decrease in the number of gauche isomers was observed relative to the number of trans isomers what corresponds to an increase in the packing density of the acylic chains. Based on derived electrical properties of the supported membranes it can be concluded that Valp induces the formation of pores and other defects in the lipid films. Valp incorporated into the membrane is seriously detrimental to the bilayer stability.

Conductivity Tensor Imaging of In Vivo Human Brain and Experimental Validation Using Giant Vesicle Suspension.

Human brain mapping of low-frequency electrical conductivity tensors can realize patient-specific volume conductor models for neuroimaging and electrical stimulation. We report experimental validation and in vivo human experiments of a new electrodeless conductivity tensor imaging (CTI) method. From CTI imaging of a giant vesicle suspension using a 9.4-T MRI scanner, the relative error in the reconstructed conductivity tensor image was found to be less than 1.7% compared with the measured value using an impedance analyzer. In vivo human brain imaging experiments of five subjects were followed using a 3-T clinical MRI scanner. With the spatial resolution of 1.87 mm, the white matter conductivity showed considerably more position dependency compared with the gray matter and cerebrospinal fluid (CSF). The anisotropy ratio of the white matter was in the range of 1.96-3.25 with a mean value of 2.43, whereas that of the gray matter was in the range of 1.12-1.19 with a mean value of 1.16. The three diagonal components of the reconstructed conductivity tensors were from 0.08 to 0.27 S/m for the white matter, from 0.20 to 0.30 S/m for the gray matter, and from 1.55 to 1.82 S/m for the CSF. The reconstructed conductivity tensor images exhibited significant inter-subject variabilities in terms of frequency and position dependencies. The high-frequency and low-frequency conductivity values can quantify the total and extracellular water contents, respectively, at every pixel. Their difference can quantify the intracellular water content at every pixel. The CTI method can separately quantify the contributions of ion concentrations and mobility to the conductivity tensor.

Movement of giant lipid vesicles induced by millimeter wave radiation change when they contain magnetic nanoparticles.

Superparamagnetic iron oxide nanoparticles are used in a rapidly expanding number of research and practical applications in biotechnology and biomedicine. Recent developments in iron oxide nanoparticle design and understanding of nanoparticle membrane interactions have led to applications in magnetically triggered, liposome delivery vehicles with controlled structure. Here we study the effect of external physical stimuli-such as millimeter wave radiation-on the induced movement of giant lipid vesicles in suspension containing or not containing iron oxide maghemite (γ-Fe2O3) nanoparticles (MNPs). To increase our understanding of this phenomenon, we used a new microscope image-based analysis to reveal millimeter wave (MMW)-induced effects on the movement of the vesicles. We found that in the lipid vesicles not containing MNPs, an exposure to MMW induced collective reorientation of vesicle motion occurring at the onset of MMW switch "on." Instead, no marked changes in the movements of lipid vesicles containing MNPs were observed at the onset of first MMW switch on, but, importantly, by examining the course followed; once the vesicles are already irradiated, a directional motion of vesicles was induced. The latter vesicles were characterized by a planar motion, absence of gravitational effects, and having trajectories spanning a range of deflection angles narrower than vesicles not containing MNPs. An explanation for this observed delayed response could be attributed to the possible interaction of MNPs with components of lipid membrane that, influencing, e.g., phospholipids density and membrane stiffening, ultimately leads to change vesicle movement.

Quenching of tryptophan fluorescence in a highly scattering solution: Insights on protein localization in a lung surfactant formulation.

CHF5633 (Chiesi Farmaceutici, Italy) is a synthetic surfactant developed for respiratory distress syndrome replacement therapy in pre-term newborn infants. CHF5633 contains two phospholipids (dipalmitoylphosphatidylcholine and 1-palmitoyl-2oleoyl-sn-glycero-3-phosphoglycerol sodium salt), and peptide analogues of surfactant protein C (SP-C analogue) and surfactant protein B (SP-B analogue). Both proteins are fundamental for an optimal surfactant activity in vivo and SP-B genetic deficiency causes lethal respiratory failure after birth. Fluorescence emission of the only tryptophan residue present in SP-B analogue (SP-C analogue has none) could in principle be exploited to probe SP-B analogue conformation, localization and interaction with other components of the pharmaceutical formulation. However, the high light scattering activity of the multi-lamellar vesicles suspension characterizing the pharmaceutical surfactant formulation represents a challenge for such studies. We show here that quenching of tryptophan fluorescence and Singular Value Decomposition analysis can be used to accurately calculate and subtract background scattering. The results indicate, with respect to Trp microenvironment, a conformationally homogeneous population of SP-B. Trp is highly accessible to the water phase, suggesting a surficial localization on the membrane of phospholipid vesicles, similarly to what observed for full length SP-B in natural lung surfactant, and supporting an analogous role in protein anchoring to the lipid phase.

Enzymatic Synthesis of Highly Electroactive Oligoanilines from a p-Aminodiphenylamine/Aniline Mixture with Anionic Vesicles as Templates.

Oligoanilines with characteristic properties of the electrically conductive emeraldine salt form of polyaniline (PANI-ES) are promising molecules for various applications. A mixture of such oligoanilines can be obtained, for example, enzymatically under mild conditions from the linear aniline dimer p-aminodiphenylamine (PADPA) with hydrogen peroxide (H2O2) and low amounts of horseradish peroxidase (HRP) in an aqueous pH = 4.3 suspension of anionic vesicles formed from AOT, the sodium salt of bis(2-ethylhexyl)sulfosuccinate. However, the simultaneous formation of undesired side products containing phenazine-type units or oxygen atoms is unsatisfactory. We have found that this situation can be improved considerably by using a mixture of PADPA and aniline instead of PADPA only but otherwise nearly identical conditions. The PANI-ES-like oligoaniline products that are obtained from the PADPA and aniline mixture were not only found to have much lower contents of phenazine-type units and not contain oxygen atoms but also were shown to be more electroactive in cyclic voltammetry measurements than the PANI-ES-like products obtained from PADPA only. The AOT vesicle suspension remained stable without product precipitation during and after the entire reaction so that it could be analyzed by in situ UV/visible/near-infrared, in situ electron paramagnetic resonance, and in situ Raman spectroscopy measurements. These measurements were complemented with ex situ high-performance liquid chromatography analyses of the deprotonated and reduced products formed from mixtures of PADPA and either fully or partially deuterated aniline. On the basis of the results obtained, a reaction mechanism is proposed for explaining this improved HRP-triggered, vesicle-assisted synthesis of electroactive PANI-ES-like products. The oligomeric products obtained can be further used, without additional special workup, for example, to coat electrodes for their possible application in biosensor devices.

Influence of Specific Bacteriophage on the Level of Vesicle Formation and Morphology of Cells of Yersinia pseudotuberculosis.

Incubation of Yersinia pseudotuberculosis cells grown on a solid medium with pseudotuberculous diagnostic bacteriophage for 20 min at 37oC led to a significant decrease in the concentration of both components of the system. This effect was absent when the bacteria were grown in a fluid medium. At the same time, this incubation regimen promoted vesicle formation and typical morphological changes in bacteria grown in both surface and suspension cultures. Co-incubation of the bacteriophage with suspension of vesicles isolated from the suspension culture of Y. pseudotuberculosis grown at 10oC (but not 37oC) led to a decrease in plaque-forming activity of the bacteriophage.

Raman spectroscopy for quantification of water-to-lipid ratio in phospholipid suspensions

In this paper, a way of quantification of the water-to-lipid ratio in aqueous suspensions of phospholipid vesicles by Raman spectroscopy is suggested. We measured the ratio of Raman intensities of the OH stretching band and the symmetric CH2-stretching mode for series of water suspensions of different phosphatidylcholines with varied water-to-phospholipid ratio. It is shown that there is a simple linear relation between the ratio of Raman intensities and the water-to-lipid ratio if the number of CH2 groups in the hydrocarbon chains is taken into account. The proportionality coefficient of this relation is determined for the gel and fluid phases of phospholipids.

Montmorency tart cherry anthocyanins: dose‐dependent antioxidant activity against cholesterol oxidation

Cardiovascular disease (CVD) is a leading cause of death world‐wide, and cholesterol oxidation (CO) and cholesterol oxidation products (COPs) are widely thought to contribute to CVD by exacerbating mechanisms that produce atherosclerotic plaques. COPs not only hold atherogenic properties, but also mutagenic, carcinogenic and inflammatory properties. More recently, it has been reported that they also play an important role in the onset and progression of neurodegenerative diseases. As promoters of inflammation, cytotoxicity, and fibrogenesis, COPs drive a cycle of inflammation that increases the steady‐state level of radical and non‐radical reactive oxygen species (ROS), which in turn amplify inflammation.Strong evidence indicates that natural products derived from plants, such as anthocyanins, poses antioxidant properties and may inhibit lipid oxidation and its metabolites. Montmorency cherries (Prunus cerasus) contain polyphenols in amounts of 3000 mg/100g dry weight, because of this they could act as a natural source of strong antioxidants. Our group has fractionated and characterized different polyphenols from Montmorency tart cherry extracts. Our preliminary studies indicate that anthocyanins have potent dose‐dependent antioxidants activity against CO suggesting that they may also be efficient at reducing CO‐induced inflammation in the CVD. We explore the antioxidant properties of Montmorency tart cherry anthocyanins against CO using in vitro systems of CVD, such as biomimetic liposomes and low‐density lipoprotein (LDL). Radical‐induced oxidation was triggered by addition of AAPH radical generator at 0.5–2 mM to a large unilamellar vesicles (LUVs) suspension under physiological conditions. Different concentrations of anthocyanins (2–100 μM) were tested over 180 minutes.Results demonstrated that anthocyanins may have a dose‐dependent antioxidant activity, specifically targeting CO. UV‐Vis spectroscopy showed that an increase in the oxidation of liposomal cholesterol when AAPH was added to the system (Control). A dramatic increase in cholesterol oxidation was observed between 180 and 240 minutes. The kinetics associated with the formation of cholesterol hydroperoxides were delayed up 180 min, with subsequent decrease on the accumulation of 7‐ketocholesterol (7‐Keto) and 25‐hydroxycholesterol (25‐OH), which are biomarkers of CO and thus the cardiovascular inflammation. We speculate that specific compounds derived from Montmorency tart cherries have the ability of to penetrate lipid membrane and scavenge ROS.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Elementary processes for the entry of cell-penetrating peptides into lipid bilayer vesicles and bacterial cells

Cell-penetrating peptides (CPPs) can translocate across the plasma membrane of living eukaryotic cells and enter the cytosol without significantly affecting cell viability. Consequently, CPPs have been used for the intracellular delivery of biological cargo such as proteins and oligonucleotides. However, the mechanisms underlying the translocation of CPPs across the plasma membrane remain unclear. In this mini-review, we summarize the experimental results regarding the entry of CPPs into lipid bilayer vesicles obtained using three methods: the large unilamellar vesicle (LUV) suspension method, the giant unilamellar vesicle (GUV) suspension method, and the single GUV method. The advantages and disadvantages of these methods are also discussed. Experimental results to date clearly indicate that CPPs can translocate across lipid bilayers and enter the vesicle lumen. Three models for the mechanisms and pathways by which CPPs translocate across lipid bilayers are described: (A) through pores induced by CPPs, (B) through transient prepores, and (C) via formation of inverted micelles. Both the pathway of translocation and the efficiency of entry of CPPs depend on the lipid composition of the bilayer and the type of CPP. We also describe the interaction of CPPs with bacterial cells. Some CPPs have strong antimicrobial activities. There are two modes of action of CPPs on bacterial cells: CPPs can induce damage to the plasma membrane and thus increase permeability, or CPPs enter the cytosol of bacterial cells without damaging the plasma membrane. The information currently available on the elementary processes by which CPPs enter lipid bilayer vesicles and bacterial cells is valuable for elucidating the mechanisms of entry of CPPs into the cytosol of various eukaryotic cells.

Acetonitrile-Induced Destabilization in Liposomes

To understand the behavior of cellular interfaces, it is important to clarify the effect of chemical compounds on artificial cell membranes. In this study, an aqueous acetonitrile solution was mixed with a suspension of lipid vesicles, and the changes in vesicle behavior arising as a result of acetonitrile application were observed. The fast Fourier transformations (FFTs) of the membrane waviness/crinkliness of the vesicles were carried out, and the membrane thermal fluctuations were analyzed. The experimental results show that the addition of acetonitrile molecules enhances the fluctuation of lipid membranes. In particular, the k = 2 mode fluctuation was significantly enhanced. This finding is expected to lead us to a further understanding of the fundamental properties of living cells.