Shape Changes Of Vesicles Research Articles

Methyl-β-cyclodextrin asymmetrically extracts phospholipid from bilayers, granting tunable control over differential stress in lipid vesicles.

Lipid asymmetry - that is, a nonuniform lipid distribution between the leaflets of a bilayer - is a ubiquitous feature of biomembranes and is implicated in several cellular phenomena. Differential tension - that is, unequal lateral monolayer tensions comparing the leaflets of a bilayer- is closely associated with lipid asymmetry underlying these varied roles. Because differential tension is not directly measurable in combination with the fact that common methods to adjust this quantity grant only semi-quantitative control over it, a detailed understanding of lipid asymmetry and differential tension are impeded. To overcome these challenges, we leveraged reversible complexation of phospholipid by methyl-β-cyclodextrin (mbCD) to tune the direction and magnitude of lipid asymmetry in synthetic vesicles. Lipid asymmetry generated in our study induced (i) vesicle shape changes and (ii) gel-liquid phase coexistence in 1-component vesicles. By applying mass-action considerations to interpret our findings, we discuss how this approach provides access to phospholipid thermodynamic potentials in bilayers containing lipid asymmetry (which are coupled to the differential tension of a bilayer). Because lipid asymmetry yielded by our approach is (i) tunable and (ii) maintained over minute to hour timescales, we anticipate that this approach will be a valuable addition to the experimental toolbox for systematic investigation into the biophysical role(s) of lipid asymmetry (and differential tension).

Thermal fluctuations and osmotic stability of lipid vesicles.

Biological membranes constantly change their shape in response to external stimuli, and understanding the remodeling and stability of vesicles in heterogeneous environments is therefore of fundamental importance for a range of cellular processes. One crucial question is how vesicles respond to external osmotic stresses, imposed by differences in solute concentrations between the vesicle interior and exterior. Previous analyses of the membrane bending energy have predicted that micron-sized giant unilamellar vesicles (GUVs) should become globally deformed already for nanomolar concentration differences, in contrast to experimental findings that find deformations at much higher osmotic stresses. In this article, we analyze the mechanical stability of a spherical vesicle exposed to an external osmotic pressure in a statistical-mechanical model, including the effect of thermally excited membrane bending modes. We find that the inclusion of thermal fluctuations of the vesicle shape changes renders the vesicle deformation continuous, in contrast to the abrupt transition in the athermal picture. Crucially, however, the predicted critical pressure associated with global vesicle deformation remains the same as when thermal fluctuations are neglected, approximately six orders of magnitude smaller than the typical collapse pressure recently observed experimentally for GUVs. We conclude by discussing possible sources of this persisting dissonance between theory and experiments.

Unraveling How Cholesterol Affects Multivalency-Induced Membrane Deformation of Sub-100 nm Lipid Vesicles.

Cholesterol plays a critical role in modulating the lipid membrane properties of biological and biomimetic systems and recent attention has focused on its role in the functions of sub-100 nm lipid vesicles and lipid nanoparticles. These functions often rely on multivalent ligand-receptor interactions involving membrane attachment and dynamic shape transformations while the extent to which cholesterol can influence such interaction processes is largely unknown. To address this question, herein, we investigated the attachment of sub-100 nm lipid vesicles containing varying cholesterol fractions (0-45 mol %) to membrane-mimicking supported lipid bilayer (SLB) platforms. Biotinylated lipids and streptavidin proteins were used as model ligands and receptors, respectively, while the localized surface plasmon resonance sensing technique was employed to track vesicle attachment kinetics in combination with analytical modeling of vesicle shape changes. Across various conditions mimicking low and high multivalency, our findings revealed that cholesterol-containing vesicles could bind to receptor-functionalized membranes but underwent appreciably less multivalency-induced shape deformation than vesicles without cholesterol, which can be explained by a cholesterol-mediated increase in membrane bending rigidity. Interestingly, the extent of vesicle deformation that occurred in response to increasingly strong multivalent interactions was less pronounced for vesicles with greater cholesterol fraction. The latter trend was rationalized by taking into account the strong dependence of the membrane bending energy on the area of the vesicle-SLB contact region and such insights can aid the engineering of membrane-enveloped nanoparticles with tailored biophysical properties.

Electromechanical characterization of biomimetic membranes using electrodeformation of vesicles.

We describe a facile method to simultaneously measure the bending rigidity and capacitance of biomimetic lipid bilayers. Our approach utilizes the ellipsoidal deformation of quasi-spherical giant unilamellar vesicles induced by a uniform AC electric field. Vesicle shape depends on the electric field frequency and amplitude. Membrane bending rigidity can be obtained from the variation of the vesicle elongation on either field amplitude at fixed frequency or frequency at fixed field amplitude. Membrane capacitance is determined from the frequency at which the vesicle shape changes from prolate to oblate ellipsoid as the frequency is increased at a given fieldamplitude.

Shaping Giant Membrane Vesicles in 3D-Printed Protein Hydrogel Cages.

Giant unilamellar phospholipid vesicles are attractive starting points for constructing minimal living cells from the bottom-up. Their membranes are compatible with many physiologically functional modules and act as selective barriers, while retaining a high morphological flexibility. However, their spherical shape renders them rather inappropriate to study phenomena that are based on distinct cell shape and polarity, such as cell division. Here, a microscale device based on 3D printed protein hydrogel is introduced to induce pH-stimulated reversible shape changes in trapped vesicles without compromising their free-standing membranes. Deformations of spheres to at least twice their aspect ratio, but also toward unusual quadratic or triangular shapes can be accomplished. Mechanical force induced by the cages to phase-separated membrane vesicles can lead to spontaneous shape deformations, from the recurrent formation of dumbbells with curved necks between domains to full budding of membrane domains as separate vesicles. Moreover, shape-tunable vesicles are particularly desirable when reconstituting geometry-sensitive protein networks, such as reaction-diffusion systems. In particular, vesicle shape changes allow to switch between different modes of self-organized protein oscillations within, and thus, to influence reaction networks directly by external mechanical cues.

Isogeometric analysis for phase-field models of geometric PDEs and high-order PDEs on stationary and evolving surfaces

In this paper, we present an isogeometric analysis (IGA) for phase-field models of three different yet closely related classes of partial differential equations (PDEs): geometric PDEs, high-order PDEs on stationary surfaces, and high-order PDEs on evolving surfaces; the latter can be a coupling of the former two classes. In the context of geometric PDEs, we consider mean curvature flow and Willmore flow problems and their corresponding phase-field approximations which yield second-order and fourth-order nonlinear parabolic PDEs. Through some numerical examples, we study the convergence behavior of isogeometric analysis for these equations using the method of manufactured solutions. Moreover, we study numerically the convergence of these phase-field approximations to the sharp interface solutions. As for the high-order PDEs on stationary surfaces, we consider a model problem which is the Cahn–Hilliard equation on a unit sphere, where the surface is modeled using a diffuse-interface approach. Finally, as a model problem for high-order PDEs on evolving surfaces, we consider a phase-field model of a deforming multicomponent vesicle which couples the vesicle shape changes with the phase separation process on the vesicle surface. The model consists of two fourth-order nonlinear PDEs which their direct finite element formulation in a Galerkin framework necessitates smooth basis functions with at least global C1 continuity; a condition that can be easily satisfied using spline bases in IGA. We solve the coupled equations both in two dimensions, where the vesicle is a curve, and in three dimensions, where the vesicle is a surface. The simulation results agree with the numerical and experimental results from the literature.

Light-Controlled Membrane Mechanics and Shape Transitions of Photoswitchable Lipid Vesicles.

Giant unilamellar vesicles (GUVs) represent a versatile model system to emulate the fundamental properties and functions associated with the plasma membrane of living cells. Deformability and shape transitions of lipid vesicles are closely linked to the mechanical properties of the bilayer membrane itself and are typically difficult to control under physiological conditions. Here, we developed a protocol to form cell-sized vesicles from an azobenzene-containing phosphatidylcholine (azo-PC), which undergoes photoisomerization on irradiation with UV-A and visible light. Photoswitching within the photolipid vesicles enabled rapid and precise control of the mechanical properties of the membrane. By varying the intensity and dynamics of the optical stimulus, controlled vesicle shape changes such as budding transitions, invagination, pearling, or the formation of membrane tubes were achieved. With this system, we could mimic the morphology changes normally seen in cells, in the absence of any molecular machines associated with the cytoskeleton. Furthermore, we devised a mechanism to utilize photoswitchable lipid membranes for storing mechanical energy and then releasing it on command as locally usable work.

Transient oscillation of shape and membrane conductivity changes by field pulse-induced electroporation in nano-sized phospholipid vesicles

The results of electrooptical and conductometrical measurements on unilamellar lipid vesicles (of mean radius a = 90 nm), filled with 0.2 M NaCl solution, suspended in 0.33 M sucrose solution of 0.2 mM NaCl, and exposed to a stepwise decaying electric field (time constant τE = 154 μs) in the range 10 ≤ E0 (kV cm(-1)) ≤ 90, are analyzed in terms of cyclic changes in vesicle shape and vesicle membrane conductivity. The two peaks in the dichroitic turbidity relaxations reflect two cycles of rapid membrane electroporation and slower resealing of long-lived electropores. The field-induced changes reflect structural transitions between closed (C) and porated (P) membrane states, qualified by pores of type P1 and of type P2, respectively. The transient change in the membrane conductivity and the transient shape oscillation are based on changes in the pore density of the (larger) P2-pores along a hysteresis cycle. The P2-pore formation leads to transient net ion flows across the P2-pores and to transient changes in the membrane field. The kinetic data are numerically processed in terms of coupled structural relaxation modes. Using the torus-hole pore model, the mean inner pore radii are estimated to be r1 = 0.38 (±0.05) nm and r2 = 1.7 (±0.1) nm, respectively. The observation of a transient oscillation of membrane electroporation and of shape changes in a longer lasting external field pulse is suggestive of potential resonance enhancement, for instance, of electro-uptake by, and of electro-release of biogenic molecules from, biological cells in trains of long-lasting low-intensity voltage pulses.

Effects of the pore-forming agent nystatin on giant phospholipid vesicles

The effects of the polyene pore-forming agent nystatin were investigated on individual giant unilamellar phospholipid vesicles (GUVs), made of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), in different methanol–water solutions using phase-contrast optical microscopy. Three characteristic effects were detected in three different nystatin concentration ranges: vesicle shape changes (between 150 and 250μM); transient, nonspecific, tension pores (between 250 and 400μM); and vesicle ruptures (above 400μM). Both the appearance of the transient tension pores and the vesicle ruptures were explained as being a consequence of the formation of size-selective nystatin channels, whose membrane area density increases with the increasing nystatin concentrations. Our results also show that nystatin is able to form pores in the absence of sterols. In addition, study of the cross-interactions between nystatin and methanol revealed mutually antagonizing effects on the vesicle behavior for methanol volume fractions higher than 10%.

Interactions in two-component liposomes studied by 2D correlation spectroscopy

The effect of dipping amphiphilic ICPANs (1-Alkylaminium, N-[2-[3-[3,5-bis(1,1-dimethylethyl)-4-hydroxyphenyl]-1-oxopropoxy]ethyl]-N,N-dimethyl-, bromide) homologues, characterized by varying alkyl chain length ( n = 8, 10, 12, and 16), into large multilamellar vesicles (MLVs) of dipalmitoylphosphatidylcholine (DPPC) was studied. Attenuated total reflectance infrared (ATR-IR) spectroscopy combined with 31P-NMR enabled observing a cut-off effect for the longest homologue. By employing two-dimensional correlation spectroscopy (2DCOS) for monitoring spectral changes induced by the heating process, detailed information about structural changes was obtained. They confirmed the substantial reorganization in the structure of the interfacial region in the ICPAN-C16/DPPC vesicles compared with the shorter homologues, where mainly the alkyl chains experience significant trans-to-gauche reorganization. Absorbance changes around 1400 cm −1 assigned to the symmetric deformation mode δ sym ( +N(CH 3) 3) are a good marker of changes in vesicle shape and are sensitive to the percentage of DPPC molecules directly interacting with the surface of the ATR crystal. This study clearly demonstrates the potential of 2DCOS in investigating interactions in two-component liposomes.

Polymorphism of vesicles with multi-domain patterns

Vesicle membranes with two coexisting fluid phases can exhibit a variety of domain patterns. These patterns and the associated vesicle shapes are studied theoretically by minimization of energy functionals that depend on the membrane composition and on the material parameters of the membrane. The latter parameters are (i) the bending rigidities and (ii) the Gaussian curvature moduli of the two types of membrane domains as well as (iii) the line tension of the domain boundaries. It is shown that the interplay between these different parameters leads to stable multi-domain patterns with more than two domains for a wide range of membrane composition and material parameters. As the membrane composition or the material parameters are varied, the vesicles can undergo transitions between different patterns of membrane domains. For fixed vesicle volume, an additional domain-induced transition is observed, in which the vesicle shape changes even though the domain pattern does not. The different multi-domain patterns and vesicle shapes are summarized in terms of morphology diagrams.

Formation and Activity of Template-Assembled Receptor Signaling Complexes

Problems in membrane biology require methods to recreate the interactions between receptors and cytoplasmic signaling proteins at the membrane surface. Here, unilamellar vesicles composed of 1,2-dioleoyl-sn-glycero-3-phosphocholine and a nickel-chelating lipid were used as templates to direct the assembly of proteins from the Escherichia coli chemotaxis signaling pathway. The bacterial chemoreceptors are known to form clusters, which promote the binding of the adaptor protein (CheW) and the kinase (CheA). When CheA was incubated with vesicles, CheW, and a histidine-tagged cytoplasmic domain fragment of the aspartate chemoreceptor (CF), the kinase activity was stimulated approximately 300-fold. Activity and pull-down assays were used with dynamic light scattering and electron microscopy to characterize the protein-vesicle compositions that were correlated with the high levels of activity, which demonstrated that CF-CheW-CheA complexes on the vesicle surface were the active entities. Assembly and stimulation occurred with vesicles of different sizes and CFs in different extents of glutamine substitution (in place of glutamate) at physiologically relevant sites. An exception was the combination of sonicated vesicles with the unsubstituted CF, which displayed lower CheA activity. The lower activity was attributed to the high curvature of the sonicated vesicles and a weaker tendency of the unsubstituted CF to self-assemble. Electron micrographs of the vesicle-protein assemblies revealed that protein binding induced pronounced changes in vesicle shape, which was consistent with the introduction of positive curvature in the outer leaflet of the bilayer. Overall, vesicle-mediated template-directed assembly is shown to be an effective way to form functional complexes of membrane-associated proteins and suggests that significant changes in membrane shape can be involved in the process of transmembrane signaling.

Electric Pulses Induce Cylindrical Deformations on Giant Vesicles in Salt Solutions

In this article, we report for the first time unusual shape changes of vesicles subjected to strong electric pulses in salt solutions of low concentration. The electric field is created by two parallel electrodes between which the vesicle solution is located. Surprisingly, the vesicles assume cylindrical shapes during the pulse. These deformations are short-lived (their lifetime is ∼1ms) and occur only in the presence of salt outside the vesicles, irrespective of their content. When the solution conductivities inside and outside are the same, vesicles with square cross section are observed. Using a fast digital camera, we were able to record these deformations and study the vesicle shape dynamics. The aims of this article are to report the new vesicle morphologies and their dynamics and to provoke theoretical work in this direction.

What can we learn about the lipid vesicle structure from the small-angle neutron scattering experiment?

Small-angle neutron scattering (SANS) on the unilamellar vesicle (ULV) populations (diameter 500 and 1,000 A) in D2O was used to characterize lipid vesicles from dimyristoylphosphatidylcholine (DMPC) at three phases: gel Lbeta', ripple Pbeta' and liquid Lalpha. Parameters of vesicle populations and internal structure of the DMPC bilayer were characterized on the basis of the separated form factor (SFF) model. Vesicle shape changes from nearly spherical in the Lalpha phase to elliptical in the Pbeta' and Lbeta' phases. This is true for vesicles prepared via extrusion through pores with the diameter 500 A. Parameters of the internal bilayer structure (thickness of the membrane and the hydrophobic core, hydration and the surface area of the lipid molecule) were determined on the basis of the hydrophobic-hydrophilic (HH) approximation of neutron scattering length density across the bilayer rhox and of the step function (SF) approximation of rhox. DMPC membrane thickness in the Lalpha phase (T = 30 degrees C) demonstrates a dependence on the membrane curvature for extruded vesicles. Prepared via extrusion through 500 A diameter pores, vesicle population in the Lalpha phase has the following characteristics: average value of minor semi-axis 266 +/- 2 A, ellipse eccentricity 1.11 +/- 0.02, polydispersity 26%, thickness of the membrane 48.9 +/- 0.2 A and of the hydrophobic core 19.9 +/- 0.4 A, surface area 60.7 +/- 0.5 A2 and number of water molecules 12.8 +/- 0.3 per DMPC molecule. Vesicles prepared via extrusion through pores with the diameter 1,000 A have polydispersity of 48% and membrane thickness of 45.5 +/- 0.6 A in the Lalpha phase. SF approximation was used to describe the DMPC membrane structure in Lbeta' (T = 10 degrees C) and Pbeta' (T = 20 degrees C) phases. Extruded DMPC vesicles in D2O have membrane thickness of 49.6 +/- 0.5 A in the Lbeta' phase and 48.3 +/- 0.6 A in the Pbeta' phase. The dependence of the DMPC membrane thickness on temperature was restored from the SANS experiment.

Membrane electroporation and electromechanical deformation of vesicles and cells

Analysis of the reduced turbidity (delta T-/T0) and absorbance (delta A-/A0) relaxations of unilamellar lipid vesicles, doped with the diphenylhexatrienyl-phosphatidylcholine (beta-DPH pPC) lipids in high-voltage rectangular electrical field pulses, demonstrates that the major part of the turbidity and absorbance dichroism is caused by vesicle elongation under electric Maxwell stress. The kinetics of this electrochemomechanical shape deformation (time constants 0.1 < or = tau/microsecond < or = 3) is determined both by the entrance of water and ions into the bulk membrane phase to form local electropores, and by the faster processes of membrane stretching and smoothing of thermal undulations. Moreover, the absorbance dichroism indicates local displacements of the chromophore relative to the membrane normal in the field. The slightly slower relaxations of the chemical turbidity (delta T+/T0) and absorbance (delta A+/A0) modes are both associated with the entrance of solvent into the interface membrane/medium, caused by the alignment of the bipolar lipid head groups in one of the leaflets at the pole caps of the vesicle bilayer. In addition, (delta T+/T0) indicates changes in vesicle shape and volume. The results for lipid vesicles provide guidelines for the analysis of electroporative deformations of biological cells.

Morphological variation in Coryphidium from the Arenigian Series (Lower Ordovician) of northwestern England

Three morphotypes of Coryphidium have been identified in the Skiddaw Group of Arenigian (Early Ordovician) age in northwestern England. Lower Arenigian rocks, probably of the Tetragraptus phyllograptoides graptolite Biozone, contain Coryphidium aff. C. elegans; middle Arenigian rocks, spanning the Didymograptus deflexus, D. nitidus and the lower part of the Isograptus gibberulus graptolite biozones, have yielded Coryphidium bohemicum?; and upper Arenigian rocks, from the upper part of the I. gibberulus and the Didymograptus hirundo graptolite biozones, contain Coryphidium aff. C. bohemicum (referred to as C. bohemicum by other authors). Coryphidium can therefore be used as a guide fossil in the Arenigian of northwestern England. The three morphotypes are distinguished by vesicle shape, process shape and size, and process distribution. Vesicle shape changes from generally straight-sided in C. aff. C. elegans and C. bohemicum? to predominantly concave-sided in C. aff. C. bohemicum. The processes of C. aff. C. elegans and C. bohemicum? are comparatively long and slender whereas those of C. aff. C. bohemicum are relatively short and stout. The processes appear to be distributed randomly on C. aff. C. elegans whereas those of C. bohemicum? and C. aff. C. bohemicum tend to be concentrated at the corners of the vesicle.

Polymer-induced shape changes and capping in giant liposomes

We studied the coupling of a polymeric amphiphile undergoing a transition from an expanded to a collapsed state of the hydrophilic polymer chain upon heating above 29°C with free and solid supported lipid bilayers. Lateral diffusion measurements and studies of vesicle shape-changes confirmed previous calorimetric studies, suggesting that the amphiphile remains incorporated in the giant vesicles and the supported bilayer during the transition. The two-dimensional hydrodynamic radii of the amphiphile in the expanded and collapsed state was measured by making use of the strong quadratic dependence of the diffusion coefficient on the radius of the diffusant ( D ∞ r −2) due to the frictional coupling between the bilayer and the supporting substrate. The hydrodynamic radius changed by a factor of 3 during, the transition from the expanded to the collapsed state. Shape-changes of multilamellar and thin-walled vesicles during the transition were observed by phase contrast microscopy. In the expanded state of the polymer, the vesicles exhibit a smooth contour, although non-trivial and non-symmetric shapes, were found. Heating over the collapse transition induces pronounced budding of the thin-walled vesicles, while the multilamellar liposomes exhibit only a transient formation of long wavelength protrusions. The shape-changes are explained in terms of the coupling between phase separation and induced curvature.

On shape transformations and shape fluctuations of cellular compartments and vesicles

We discuss the shape formation and shape transitions of simple bilayer vesicles in context with their role in biology. In the first part several classes of shape changes of vesicles of one lipid component are described and it is shown that these can be explained in terms of the bending energy concept in particular augmented by the bilayer coupling hypothesis. In the second part shape changes and vesicle fission of vesicles composed of membranes of lipid mixtures are reported. These are explained in terms of coupling between local curvature and phase separation.

Morphological effects of osmolarity on purified noradrenergic vesicles.

Large dense core vesicles (LDV) were purified from bovine splenic nerve homogenates by the sucrose-D2O density gradient method. Vesicles were subjected to a 50% increase and decrease in osmolarity from the control 330 mosmol 1(-1) by adjusting sucrose or potassium phosphate buffer during pre- and/or postfixation. Control vesicles with a mean diameter of 717 A readily swelled to approximately 1050 A and shrunk to approximately 600 A in the hypotonic and hypertonic media, respectively, with either sucrose or phosphate buffer. The dense core responded similarly but to a lesser degree. Prefixation in glutaraldehyde had little effect on vesicle sensitivity to subsequent tonicity change, not did the fixative per se exert an obvious osmotic effect. Thus, final vesicle size was largely determined by the OsO4 postfixation medium and principally by the vehicle rather than the fixative. In controls there was a mixture of spherical to oblate vesicles mostly filled with an electron-dense matrix. Upon swelling, more vesicles became spherical and nearly all had a prominent translucent halo between core and membrane. Upon shrinking, more vesicles became oblate, the halo was obliterated and the electron-density of the matrix increased. Frequency distributions of vesicle diameters at different tonicity clearly indicated that the diameter of LDV could overlap the 400-500 A range characteristic of small dense core vesicles; however, there was no suggestion of a population of the latter in the purified LDV fraction. Implications are discussed concerning the biochemical and morphological identification of 'light' and 'heavy' density peaks of noradrenaline and dopamine beta-hydroxylase from mixed vesicle populations and the possible relevance of changes in vesicle shape to a functional state in situ.