Lipid Vesicles Research Articles

HylleraasMD: A Domain Decomposition-Based Hybrid Particle-Field Software for Multiscale Simulations of Soft Matter

We present HylleraasMD (HyMD), a comprehensive implementation of the recently proposed Hamiltonian formulation of hybrid particle-field molecular dynamics. The methodology is based on a tunable, grid-independent length-scale of coarse graining, obtained by filtering particle densities in reciprocal space. This enables systematic convergence of energies and forces by grid refinement, also eliminating nonphysical force aliasing. Separating the time integration of fast modes associated with internal molecular motion from slow modes associated with their density fields, we enable the first time-reversible, energy-conserving hybrid particle-field simulations. HyMD comprises the optional use of explicit electrostatics, which, in this formalism, corresponds to the long-range potential in particle-mesh Ewald. We demonstrate the ability of HyMD to perform simulations in the microcanonical and canonical ensembles with a series of test cases, comprising lipid bilayers and vesicles, surfactant micelles, and polypeptide chains, comparing our results to established literature. An on-the-fly increase of the characteristic coarse-grain length significantly speeds up dynamics, accelerating self-diffusion and leading to expedited aggregation. Exploiting this acceleration, we find that the time scales involved in the self-assembly of polymeric structures can lie in the tens to hundreds of picoseconds instead of the multimicrosecond regime observed with comparable coarse-grained models.

Fluorescent macrocycle-dye-anchor conjugates for sensing phospholipids in biomembranes

Negatively charged phospholipids can serve as biomarkers for a number of biological processes allowing detection of apoptosis in animal cells, drug-induced phospholipidosis as well as detection of bacteria. Selective detection of phospholipids thus represents an important but challenging goal. The development of fluorescent probes capable of selective recognition of specific phospholipids can be a promising solution to the problem. Along this line, we have developed the macrocycle-dye-anchor conjugates, in which the macrocycle recognizes the phosphate residue, hydrocarbon anchors ensure hydrophobic interaction with the membrane, and environment-sensitive fluorescent dye functions as a reporter unit. To the best of our knowledge, this is the first report of synthetic fluorescent probes for specific phospholipids using the macrocycle-dye-anchor concept. We have studied the effect of the structure of the anchor and the macrocycle on the resulting fluorescent response in the presence of two types of lipid vesicles: phosphatidylcholine as a model of healthy cells and phosphatidylcholine and phosphatidylserine (1:1), mimicking apoptotic cells. The properties of the obtained compounds were analyzed in various model media and in HeLa cells. The response of fluorescent probes has been found to be strongly dependent on the macrocycle structure, length of the anchor, nature of the buffer, and pH. The probes showed significant fluorescence enhancement in apoptotic cells, thus providing the proof of concept for the macrocycle-dye-anchor design.

Cell‐Sized Lipid Vesicles as Artificial Antigen‐Presenting Cells for Antigen‐Specific T Cell Activation (Adv. Healthcare Mater. 12/2023)

Immunotherapy This image depicts cell-mimicking artificial antigen-presenting cells (aAPCs, pink) and biological T cells (blue). In article 2203163 by Anshu Agrawal, Abraham P. Lee, and co-workers, aAPCs are produced using a microfluidic device, which enables facile and stable double emulsion droplet generation. By recapitulating the properties of a cell, namely, size, fluidity, and surface proteins, aAPCs are able to engage with T cells, forming immune synapses. Multiple ligands on aAPCs can engage with the receptors on T cells due to the high surface fluidity of the lipid bilayer. The size of the aAPC (≈20 μm diameter) also contributes to an adequate force that is essential for T cell activation. Experimental results suggest that T cells can tightly bind with the aAPCs and maximize their contact area. After interacting with aAPCs, T cells are activated and subsequently proliferate in large numbers. The background shows a wide spectrum of aAPCs, T cells, and most importantly, aAPC-T cell pairs joint by immune synapses, as is observed in this research.

Heterozygosity for Crohn’s Disease Risk Allele of Atg16L1 Protects from Salmonella Infection

Abstract Autophagy is an evolutionarily conserved pathway for the formation and trafficking of lipid vesicles in response to stress, including inflammation and microbial infection. A missense variant of the essential autophagy gene Atg16L1 (rs2241880, T300A) is carried by a majority of the human population and is linked to Crohn’s Disease (CD). Previous studies found that T300A enhances proteolytic cleavage and degradation of Atg16L1, leading to dysregulated cytokine expression and impaired control of microbial infection in knock-in mice. However, these studies have focused on homozygous animals while the plurality of the human population is heterozygous for the allele. Here we demonstrate that heterozygosity for T300A protects mice from Salmonella infection. We find the variant allele results in altered complex formation independent of cleavage, allowing for enhanced inflammasome activation but impaired conjugation of Atg8 to single membranes (CASM). In vivo, one copy of T300A is sufficient to elevate the production of inflammatory cytokines, which is required for protection from Salmonella infection. Meanwhile, mice deficient in CASM genes are susceptible to Salmonella infection and this pathway is shown to be impaired in homozygous but not heterozygous mice. Finally, we demonstrate that heterozygosity in human macrophages also results in elevated cytokine expression, while maintaining functional non-canonical autophagy. These results clarify the altered autophagy function of the CD risk allele Atg16L1T300A and suggest it can be beneficial to heterozygous carriers, providing a potential explanation for its prevalence in the human population. NIH R01 AI121244 F31 HL149238

Intact GR dimerization is critical for restraining plasma ACTH levels during chronic psychosocial stress

Male C57BL/6N mice exposed to the chronic subordinate colony housing (CSC; 19 days) paradigm, a preclinically validated model of chronic psychosocial stress, are characterized by unaffected basal morning plasma corticosterone (CORT) concentrations despite adrenal and pituitary hyperplasia and increased adrenocorticotropic hormone (ACTH) plasma concentrations, compared with single-housed control (SHC) mice. However, as CSC mice are still able to show an increased CORT secretion towards novel heterotypic stressors, these effects might reflect an adaptation rather than a functional breakdown of general hypothalamus-pituitary-adrenal (HPA) axis functionality. In the present study we used male mice of a genetically modified mouse line, to investigate whether genetically-driven ACTH overexpression compromises adaptational processes occurring at the level of the adrenals during CSC exposure. Experimental mice carried a point mutation in the DNA binding domain of the glucocorticoid (GC) receptor (GR), attenuating dimerization of GR (GRdim), resulting in a congenially compromised negative feedback inhibition at the level of the pituitary.In line with previous studies, CSC mice in both the wild type (WT; GR+/+) and GRdim group developed adrenal enlargement. Moreover, compared with respective SHC and WT mice, CSC GRdim mice show increased basal morning plasma ACTH and CORT concentrations. Quantitative polymerase chain reaction (qPCR) analysis revealed neither a genotype effect, nor a CSC effect on pituitary mRNA expression of the ACTH precursor proopiomelanocortin (POMC). Finally, CSC increased anxiety-related behavior, active coping and splenocyte in vitro (re)activity in both WT and GRdim mice, while a CSC-induced increase in adrenal lipid vesicles and splenic GC resistance was detectable only in WT mice. Of note, lipopolysaccharide (LPS)-stimulated splenocytes of GRdim mice were resistant to the inhibitory effects of CORT.Together our findings support the hypothesis that pituitary ACTH protein concentration is negatively controlled by GR dimerization under conditions of chronic psychosocial stress, while POMC gene transcription is not dependent on intact GR dimerization under both basal and chronic stress conditions. Finally, our data suggest that adrenal adaptations during chronic psychosocial stress (i.e., ACTH desensitization), aiming at the prevention of prolonged hypercorticism, are protective only to a certain threshold of plasma ACTH levels.

Development and quality assessment of glutathione tripeptide loaded niosome containing carbopol emulgels as nanocosmeceutical formulations

This study focuses on the preparation, physicopharmaceutical and mechanical characterization of reduced glutathione tripeptide loaded niosome containing emulgels as a novel nanocosmeceutical product. Prepared emulgel formulations were mainly composed of oily phase containing different lipids such as glycerine dibehenate, cetyl alcohol, cetearyl alcohol, etc., and aqueous phase containing Carbopol934® as gelling agent. Niosomal lipidic vesicles prepared from Span 60 and cholesterol were subsequently incorporated into optimum emulgel formulations. The pH, viscosity, and textural/mechanical properties of emulgels were examined before and after the incorporation of niosomes. The viscoelasticity and morphological characterization were performed on the final formulation before the packed formulation's microbiological stability test. The hardness and compressibility results ensured easy removal of the emulgel from the container. Due to the carboxyl groups of Carbopol934®, moderate adhesiveness with good cohesiveness was achieved. The rheological characteristics of the emulgels were estimated by oscillatory testing and the data fitted with the Herschel-Bulkley model. Thus, the viscoelastic properties and shear-thinning flow of emulgels were demonstrated. The final formulation was microbiologically stable, and pathogens or skin-irritating allergens were not detected. An anti-aging cosmeceutical preparation containing glutathione tripeptide loaded lipid-based niosome dispersion, suitable for topical use due to its textural and viscosity properties, was successfully produced.

Using Holographic Microscopy To Measure the Effect of Confinement on Crowding Agents in Lipid Vesicles.

The hydrodynamic effects of macromolecular crowding inside cells are often studied in vitro by using polymers as crowding agents. Confinement of polymers inside cell-sized droplets has been shown to affect the diffusion of small molecules. Here we develop a method, based on digital holographic microscopy, to measure the diffusion of polystyrene microspheres that are confined within lipid vesicles containing a high concentration of solute. We apply the method to three solutes of varying complexity: sucrose, dextran, and PEG, prepared at ∼7 % (w/w). We find that diffusion inside and outside the vesicles is the same when the solute is sucrose or dextran that is prepared below the critical overlap concentration. For poly(ethylene glycol), which is present at a concentration higher than the critical overlap concentration, the diffusion of microspheres inside vesicles is slower, hinting at the potential effects of confinement on crowding agents.

Key Role of Choline Head Groups in Large Unilamellar Phospholipid Vesicles for the Interaction with and Rupture by Silica Nanoparticles.

For highly abundant silica nanomaterials, detrimental effects on proteins and phospholipids are postulated as critical molecular initiating events that involve hydrogen-bonding, hydrophobic, and/or hydrophilic interactions. Here, large unilamellar vesicles with various well-defined phospholipid compositions are used as biomimetic models to recapitulate membranolysis, a process known to be induced by silica nanoparticles in human cells. Differential analysis of the dominant phospholipids determined in membranes of alveolar lung epithelial cells demonstrates that the quaternary ammonium head groups of phosphatidylcholine and sphingomyelin play a critical and dose-dependent role in vesicle binding and rupture by amorphous colloidal silica nanoparticles. Surface modification by either protein adsorption or by covalent coupling of carboxyl groups suppresses the disintegration of these lipid vesicles, as well as membranolysis in human A549 lung epithelial cells by the silica nanoparticles. Furthermore, molecular modeling suggests a preferential affinity of silanol groups for choline head groups, which is also modulated by the pH value. Biomimetic lipid vesicles can thus be used to better understand specific phospholipid-nanoparticle interactions at the molecular level to support the rational design of safe advanced materials.

Quantifying the tuneable interactions between colloid supported lipid bilayers

Colloid supported lipid bilayers (CSLBs) are formed via the rupture and fusion of lipid vesicles to coat spherical colloidal particles. CSLBs are an emerging vector for the controlled self-assembly of colloids due to the ability to include additives into the bilayer, which influence the (a)specific interactions between particles. To evaluate the specificity of CSLB assembly, first a fundamental study on the tunability of the colloidal interaction and resulting colloidal stability of CSLBs without specific interactions is reported here. It was found that both fluid and gel CSLBs showed significant clustering and attraction, while the addition of steric stabilizers induced a profound increase in stability. The interactions were rendered attractive again by the introduction of depletion forces via the addition of free non-adsorbing polymers. The compositions of fluid and gel CSLBs with 5% membrane stabiliser were concluded to be optimal for further studies where both colloidal stability, and contrasting membrane fluidity are required. These experimental findings were confirmed semi-quantitatively by predictions using numerical self-consistent mean-field theory lattice computations.

A Photoswitchable Solvatochromic Dye for Probing Membrane Ordering by RESOLFT Super-resolution Microscopy.

A switchable solvatochromic fluorescent dyad can be used to map ordering of lipids in vesicle membranes at a resolution better than the diffraction limit. Combining a Nile Red fluorophore with a photochromic spironaphthoxazine quencher allows the fluorescence to be controlled using visible light, via photoswitching and FRET quenching. Synthetic lipid vesicles of varying composition were imaged with an average 2.5-fold resolution enhancement, compared to the confocal images. Ratiometric detection was used to probe the membrane polarity, and domains of different lipid ordering were distinguished within the same membrane.

Lipophilicity Modulations by Fluorination Correlate with Membrane Partitioning

AbstractBioactive compounds generally need to cross membranes to arrive at their site of action. The octanol‐water partition coefficient (lipophilicity, logPOW) has proven to be an excellent proxy for membrane permeability. In modern drug discovery, logPOW and bioactivity are optimized simultaneously, for which fluorination is one of the relevant strategies. The question arises as to which extent the often subtle logP modifications resulting from different aliphatic fluorine‐motif introductions also lead to concomitant membrane permeability changes, given the difference in molecular environment between octanol and (anisotropic) membranes. It was found that for a given compound class, there is excellent correlation between logPOW values with the corresponding membrane molar partitioning coefficients (logKp); a study enabled by novel solid‐state 19F NMR MAS methodology using lipid vesicles. Our results show that the factors that cause modulation of octanol‐water partition coefficients similarly affect membrane permeability.

Lipophilicity Modulations by Fluorination Correlate with Membrane Partitioning.

Bioactive compounds generally need to cross membranes to arrive at their site of action. The octanol-water partition coefficient (lipophilicity, logPOW ) has proven to be an excellent proxy for membrane permeability. In modern drug discovery, logPOW and bioactivity are optimized simultaneously, for which fluorination is one of the relevant strategies. The question arises as to which extent the often subtle logP modifications resulting from different aliphatic fluorine-motif introductions also lead to concomitant membrane permeability changes, given the difference in molecular environment between octanol and (anisotropic) membranes. It was found that for a given compound class, there is excellent correlation between logPOW values with the corresponding membrane molar partitioning coefficients (logKp ); a study enabled by novel solid-state 19 F NMR MAS methodology using lipid vesicles. Our results show that the factors that cause modulation of octanol-water partition coefficients similarly affect membrane permeability.

Lipid phosphatase SAC1 suppresses hepatitis B virus replication through promoting autophagic degradation of virions

Phosphatidylinositol lipids play vital roles in lipid signal transduction, membrane recognition, vesicle transport, and viral replication. Previous studies have revealed that SAC1-like phosphatidylinositol phosphatase (SACM1L/SAC1), which uses phosphatidylinositol-4-phosphate (PI4P) as its substrate, greatly affects the replication of certain bacteria and viruses in vitro. However, it remains unclear whether and how SAC1 modulates hepatitis B virus (HBV) replication in vitro and in vivo. In the present study, we observed that SAC1 silencing significantly increased HBV DNA replication, subviral particle (SVP) expression, and secretion of HBV virions, whereas SAC1 overexpression exerted the opposite effects. Moreover, SAC1 overexpression inhibited HBV DNA replication and SVP expression in a hydrodynamic injection-based HBV-persistent replicating mouse model. Mechanistically, SAC1 silencing increased the number of HBV-containing autophagosomes as well as PI4P levels on the autophagosome membrane. Moreover, SAC1 silencing blocked autophagosome-lysosome fusion by inhibiting the interaction between synaptosomal-associated protein 29 and vesicle-associated membrane protein 8. Collectively, our data indicate that SAC1 significantly inhibits HBV replication by promoting the autophagic degradation of HBV virions. Our findings support that SAC1-mediated phospholipid metabolism greatly modulates certain steps of the HBV life-cycle and provide a new theoretical basis for antiviral therapy.

Function Investigations and Applications of Membrane Proteins on Artificial Lipid Membranes.

Membrane proteins play an important role in key cellular functions, such as signal transduction, apoptosis, and metabolism. Therefore, structural and functional studies of these proteins are essential in fields such as fundamental biology, medical science, pharmacology, biotechnology, and bioengineering. However, observing the precise elemental reactions and structures of membrane proteins is difficult, despite their functioning through interactions with various biomolecules in living cells. To investigate these properties, methodologies have been developed to study the functions of membrane proteins that have been purified from biological cells. In this paper, we introduce various methods for creating liposomes or lipid vesicles, from conventional to recent approaches, as well as techniques for reconstituting membrane proteins into artificial membranes. We also cover the different types of artificial membranes that can be used to observe the functions of reconstituted membrane proteins, including their structure, number of transmembrane domains, and functional type. Finally, we discuss the reconstitution of membrane proteins using a cell-free synthesis system and the reconstitution and function of multiple membrane proteins.

Establishment of an Antiplasmodial Vaccine Based on PfRH5-Encoding RNA Replicons Stabilized by Cationic Liposomes.

Nucleic acid-based vaccines have been studied for the past four decades, but the approval of the first messenger RNA (mRNA) vaccines during the COVID-19 pandemic opened renewed perspectives for the development of similar vaccines against different infectious diseases. Presently available mRNA vaccines are based on non-replicative mRNA, which contains modified nucleosides encased in lipid vesicles, allowing for entry into the host cell cytoplasm, and reducing inflammatory reactions. An alternative immunization strategy employs self-amplifying mRNA (samRNA) derived from alphaviruses, but lacks viral structural genes. Once incorporated into ionizable lipid shells, these vaccines lead to enhanced gene expression, and lower mRNA doses are required to induce protective immune responses. In the present study, we tested a samRNA vaccine formulation based on the SP6 Venezuelan equine encephalitis (VEE) vector incorporated into cationic liposomes (dimethyldioctadecyl ammonium bromide and a cholesterol derivative). Three vaccines were generated that encoded two reporter genes (GFP and nanoLuc) and the Plasmodium falciparum reticulocyte binding protein homologue 5 (PfRH5). Transfection assays were performed using Vero and HEK293T cells, and the mice were immunized via the intradermal route using a tattooing device. The liposome-replicon complexes showed high transfection efficiencies with in vitro cultured cells, whereas tattooing immunization with GFP-encoding replicons demonstrated gene expression in mouse skin up to 48 h after immunization. Mice immunized with liposomal PfRH5-encoding RNA replicons elicited antibodies that recognized the native protein expressed in P. falciparum schizont extracts, and inhibited the growth of the parasite in vitro. Intradermal delivery of cationic lipid-encapsulated samRNA constructs is a feasible approach for developing future malaria vaccines.

Cubosomes in Drug Delivery-A Comprehensive Review on Its Structural Components, Preparation Techniques and Therapeutic Applications.

Cubosomes are lipid vesicles that are comparable to vesicular systems like liposomes. Cubosomes are created with certain amphiphilic lipids in the presence of a suitable stabiliser. Since its discovery and designation, self-assembled cubosomes as active drug delivery vehicles have drawn much attention and interest. Oral, ocular, transdermal, and chemotherapeutic are just a few of the drug delivery methods in which they are used. Cubosomes show tremendous potential in drug nanoformulations for cancer therapeutics because of their prospective advantages, which include high drug dispersal due to the structure of the cubic, large surface area, a relatively simple manufacturing process, biodegradability, ability to encapsulate hydrophobic, hydrophilic, and amphiphilic compounds, targeted and controlled release of bioactive agents, and biodegradability of lipids. The most typical technique of preparation is the simple emulsification of a monoglyceride with a polymer, followed by sonication and homogenisation. Top-down and bottom-up are two different sorts of preparation techniques. This review will critically analyse the composition, preparation techniques, drug encapsulation approaches, drug loading, release mechanism and applications relevant to cubosomes. Furthermore, the challenges faced in optimising various parameters to enhance the loading capacities and future potentialities are also addressed.

Insulin Activation Mediated by Uptake Mechanisms: A Comparison of the Behavior between Polymer Nanoparticles and Extracellular Vesicles in 3D Liver Tissues.

In this work, we compare the role of two different uptake mechanisms in the effectiveness of a nanoformulated drug, specifically insulin. Insulin is activated by interacting with insulin receptors exposed on the liver cell membrane that triggers the uptake and storage of glucose. To prove that the uptake mechanism of a delivery system can interfere directly with the effectiveness of the delivered drug, two extremely different delivery systems are tested. In detail, hydrogel-based NPs (cHANPs) and natural lipid vesicles (EVs) encapsulating insulin are used to trigger the activation of this hormone in 3D liver microtissues (μTs) based on their different uptake mechanisms. Results demonstrated that the fusion mechanism of Ins-EVs mediates faster and more pronounced insulin activation with respect to the endocytic mechanism of Ins-cHANPs. Indeed, the fusion causes an increased reduction in glucose concentration in the culture medium EV-treated l-μTs with respect to free insulin-treated tissues. The same effect is not observed for Ins-cHANPs that, taken up by endocytosis, can only equal the reduction in glucose concentration produced by free insulin in 48 h. Overall, these results demonstrate that the effectiveness of nanoformulated drugs depends on the identity they acquire in the biological context (biological identity). Indeed, the nanoparticle (NP) biological identity, such as the uptake mechanism, triggers a unique set of nano-bio-interactions that is ultimately responsible for their fate both in the extracellular and intracellular compartments.

The interplay of geometry and coarsening in multicomponent lipid vesicles under the influence of hydrodynamics

We consider the impact of surface hydrodynamics on the interplay between membrane curvature and lipid composition in coarsening processes on model systems for biomembranes. This includes the influence on scaling laws and equilibrium configurations, which are investigated by computational studies of a surface two-phase flow problem with additional phase-dependent bending terms. These additional terms geometrically favor specific configurations. We find that the effect of hydrodynamics strongly depends on the composition. In situations where the composition allows a realization of a geometrically favored configuration, hydrodynamics enhances the evolution toward this configuration. We restrict our model and numerics to stationary surfaces of varying curvature and validate the numerical approach with various benchmark problems and convergence studies.

N,N,N-Trimethyl chitosan as a permeation enhancer for inhalation drug delivery: Interaction with a model pulmonary surfactant

N,N,N-Trimethyl chitosan (TMC), a biocompatible and biodegradable derivative of chitosan, is currently used as a permeation enhancer to increase the translocation of drugs to the bloodstream in the lungs. This article discusses the effect of TMC on a mimetic pulmonary surfactant, Curosurf®, a low-viscosity lipid formulation administered to preterm infants with acute respiratory distress syndrome. Curosurf® exhibits a strong interaction with TMC, resulting in the formation of aggregates at electrostatic charge stoichiometry. At nanoscale, Curosurf® undergoes a profound reorganization of its lipid vesicles in terms of size and lamellarity. The initial micron-sized vesicles (average size 4.8 μm) give way to a froth-like network of unilamellar vesicles about 300 nm in size. Under such conditions, neutralization of the cationic charges by pulmonary surfactant may inhibit TMC permeation enhancer capacity, especially as electrostatic charge complexation is found at low TMC content. The permeation properties of pulmonary surfactant-neutralized TMC should then be evaluated for its applicability as a permeation enhancer for inhalation in the alveolar region.

Accurate Reduced Models for the pH Oscillations in the Urea–Urease Reaction Confined to Giant Lipid Vesicles

This theoretical study concerns a pH oscillator based on the urea-urease reaction confined to giant lipid vesicles. Under suitable conditions, differential transport of urea and hydrogen ion across the unilamellar vesicle membrane periodically resets the pH clock that switches the system from acid to basic, resulting in self-sustained oscillations. We analyze the structure of the phase flow and of the limit cycle, which controls the dynamics for giant vesicles and dominates the pronouncedly stochastic oscillations in small vesicles of submicrometer size. To this end, we derive reduced models, which are amenable to analytic treatments that are complemented by numerical solutions, and obtain the period and amplitude of the oscillations as well as the parameter domain, where oscillatory behavior persists. We show that the accuracy of these predictions is highly sensitive to the employed reduction scheme. In particular, we suggest an accurate two-variable model and show its equivalence to a three-variable model that admits an interpretation in terms of a chemical reaction network. The faithful modeling of a single pH oscillator appears crucial for rationalizing experiments and understanding communication of vesicles and synchronization of rhythms.