Model Cell Membranes Research Articles

Selective Glycan Presentation in Liquid‐Ordered or ‐Disordered Membrane Phases and its Effect on Lectin Binding

Glycan‐protein interactions play a key role in various biological processes from fertilization to infections. Many of these interactions take place at the glycocalyx – a heavily glycosylated layer at the cell surface. Despite its significance, studying the glycocalyx remains challenging due to its complex, dynamic, and heterogeneous nature. This study introduces a glycocalyx model allowing for the first time to control spatial organization and heterogeneity of the glycan moieties. Glycan‐mimetics with lipid‐moieties that partition into either liquid‐ordered (Lo, lipid rafts) or liquid‐disordered (Ld) phases of giant unilamellar vesicles (GUVs), which serve as simplified cell membrane models micking lipid rafts, are developed. This phase‐specific allocation allows controlled placement of glycan motifs in distinct membrane environments, creating heteromultivalent systems that replicate the natural glycocalyx's complexity. We show that phase localization of glycan mimetics significantly influences recruitment of protein receptors to the membrane. Glycan‐conjugates in the ordered phase demonstrate enhanced lectin binding, supporting the idea that raft‐like domains facilitate stronger receptor interactions. This study provides a platform for systematically investigating spatial and dynamic presentation of glycans in biological systems and presents the first experimental evidence that glycan accumulation in lipid rafts enhances receptor binding affinity, offering deeper insights into the glycocalyx's functional mechanisms.

Selective Glycan Presentation in Liquid-Ordered or -Disordered Membrane Phases and its Effect on Lectin Binding.

Glycan-protein interactions play a key role in various biological processes from fertilization to infections. Many of these interactions take place at the glycocalyx - a heavily glycosylated layer at the cell surface. Despite its significance, studying the glycocalyx remains challenging due to its complex, dynamic, and heterogeneous nature. This study introduces a glycocalyx model allowing for the first time to control spatial organization and heterogeneity of the glycan moieties. Glycan-mimetics with lipid-moieties that partition into either liquid-ordered (Lo, lipid rafts) or liquid-disordered (Ld) phases of giant unilamellar vesicles (GUVs), which serve as simplified cell membrane models micking lipid rafts, are developed. This phase-specific allocation allows controlled placement of glycan motifs in distinct membrane environments, creating heteromultivalent systems that replicate the natural glycocalyx's complexity. We show that phase localization of glycan mimetics significantly influences recruitment of protein receptors to the membrane. Glycan-conjugates in the ordered phase demonstrate enhanced lectin binding, supporting the idea that raft-like domains facilitate stronger receptor interactions. This study provides a platform for systematically investigating spatial and dynamic presentation of glycans in biological systems and presents the first experimental evidence that glycan accumulation in lipid rafts enhances receptor binding affinity, offering deeper insights into the glycocalyx's functional mechanisms.

A paradigm for natural eutectic solvents based on fatty acids: Molecular interactions and toxicological considerations

In this work, we present experimental and molecular modeling results on archetypal hydrophobic natural deep eutectic solvents (NADES) based on fatty acids (octanoic and dodecanoic acid) and menthol, a representative monoterpenoid. Our goal is to provide a multiscale characterization to enhance the understanding of this field by studying these selected archetypical mixtures. We examine their liquid state properties, intermolecular forces, nanoscopic arrangements, toxicity, and environmental impact.The computational study integrates quantum chemistry, molecular dynamics (both all-atom and coarse-grained approaches), and thermodynamic modeling (COSMO-RS approach) to analyze the fluids and their interactions with biological entities, such as proteins and plasma membranes. The experimental characterization focuses on elucidating intermolecular interactions and liquid phase dynamics using NMR spectroscopy, visible and UV Resonance Raman spectroscopy (UVRR). Notably, this is the first report of UVRR data on NADES.Additionally, we simulate the effect of the molecular moieties forming the solvents on biological targets—specifically, protein and cell membrane models –. This in silico analysis aims to rationalize and predict their potential toxicity.Overall, our experimental findings and in silico simulations contribute to a deeper understanding of these novel solvents in terms of their network of interactions. Additionally, they highlight the potential impact on biological targets, providing new data to accurately define the eco-friendliness of type V DES and their suitability as sustainable alternatives to traditional molecular solvents.

Probing the Influence of Hydrophobicity of Modified Gold Nanoparticles in Modulating the Lipid Surface Behavior Using Vibrational Sum Frequency Generation Spectroscopy.

A deep understanding of how the surface modifications of nanoparticles impact their interactions with cell membranes is vital for advancing safe and effective biomedical applications. Among the pivotal factors governing these interactions, the hydrophobicity of nanoparticles plays a crucial role, predominantly driven by the hydrophobic interactions with the cell membrane. Herein, we study the influence of the hydrophobic alkyl chain length of thiol-capped gold nanoparticles (GNPs) on lipid surfaces with the help of vibrational sum frequency generation spectroscopy. We have utilized the zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayer as a representative model of cell membranes on the water surface. Our findings revealed that GNPs capped with the thiol ligand having a shorter alkyl chain such as heptanethiol (HT, C7) show minimal changes in the C-H stretching vibrations while interacting with the lipid monolayer. These observations could be attributed to the perturbation of the lipid chain due to hydrophobic-hydrophobic interactions between the alkyl chain of thiol-capped GNPs and the hydrophobic group of the lipid membrane or simply by the adsorption of GNPs at the interface without disrupting the monolayer structure. However, with increasing the chain length of thiol-capped GNPs from decanethiol (DDT, C10) to octadecanethiol (ODT, C18), the extent of spectral change in the C-H stretching vibration is increased. The controlled experiment performed with the deuterated lipids conforms that the changes observed in the C-H stretching vibration after adding HT (C7) GNPs are only because of their presence in the surface without altering the monolayer structure. However, in the case of DT (C10) and DDT (C12) GNPs, the strong hydrophobic interactions between the monolayer and the alkyl chain of the thiol-capped GNPs result in the increased orientational order of the monolayer. Moreover, in the case of ODT (C18) GNPs, the very long alkyl chain induces pronounced perturbations in the monolayer structure with net disordering of the monolayer. These observations are further supported by the spectral changes observed in the O-H vibration of the interfacial water molecules. Our findings reveal the crucial role of the hydrophobic nature of GNPs in influencing the interface. Understanding these effects is crucial for drug delivery applications and improving the stability and effectiveness of lipid-based systems.

Evaluation of the Protective Role of Vitamin E against ROS-Driven Lipid Oxidation in Model Cell Membranes.

Reactive oxygen species (ROS) are chemically reactive oxygen-containing compounds generated by various factors in the body. Antioxidants mitigate the damaging effects of ROS by playing a critical role in regulating redox balance and signaling. In this study, the interplay between reactive oxygen species (ROS) and antioxidants in the context of lipid dynamics were investigated. The interaction between hydrogen peroxide (H2O2) as an ROS and vitamin E (α-tocopherol) as an antioxidant was examined. Model membranes containing both saturated and unsaturated lipids served as experimental platforms to investigate the influence of H2O2 on phospholipid unsaturation and the role of antioxidants in this process. The results demonstrated that H2O2 has a negative effect on membrane stability and disrupts the lipid membrane structure, whereas the presence of antioxidants protects the lipid membrane from the detrimental effects of ROS. The model membranes used here are a useful tool for understanding ROS-antioxidant interactions at the molecular level in vitro.

Probing the relevance of synergistic lipid membrane disruption to the eye irritation of binary mixed nonionic surfactants

Nonionic surfactant aerosols play a crucial role in many industries, but they can cause acute irritation to users’ eyes during spraying. This cytotoxic process is associated with corneal cell necrosis causing cell membrane disruption. Industrial grade surfactants are typically polydisperse mixtures described by their nominal chemical structure but how the polydispersity affects their interactions with cell membrane, remains largely unexplored. A better understanding could benefit product formulations to maximise their efficiency whilst minimising their toxicity to the users. In this study, poly-oxyethylene glycol monododecyl ethers (C12E4, C12E23) were used to form ideal binary surfactant mixtures. The cytotoxicities of mono and polydispersed surfactants towards human corneal epithelial cells were examined, followed by a series of biophysical characterisations of interactions between surfactants and model cell membranes. Notably, to probe the journey of individual C12E4 and C12E23 surfactant molecules across the cell membrane from a binary surfactant mixture, “two-colour” neutron reflection measurements were achieved via Hydrogen/Deuterium substitution. The relative distributions of C12E4 and C12E23 across cell membranes and their nanostructural conformations revealed a synergistic membrane-lytic ability initiated by surfactant mixing, with the more hydrophobic C12E4 exhibiting stronger membrane binding potency than the hydrophilic C12E23. The exact molar ratio of C12E4 against C12E23 in the mixture determined how the mixed surfactant interacted with the cell membrane, and how the process directly impacted cytotoxicity and eye irritation. Thus, the cytotoxicity of polydisperse surfactants is not the same as monodisperse surfactant of the same average structure. This work provides a useful basis for the assessment of surfactant mixing by balancing their efficiency and toxicity.

Advancements in Engineering Planar Model Cell Membranes: Current Techniques, Applications, and Future Perspectives.

Cell membranes are crucial elements in living organisms, serving as protective barriers and providing structural support for cells. They regulate numerous exchange and communication processes between cells and their environment, including interactions with other cells, tissues, ions, xenobiotics, and drugs. However, the complexity and heterogeneity of cell membranes-comprising two asymmetric layers with varying compositions across different cell types and states (e.g., healthy vs. diseased)-along with the challenges of manipulating real cell membranes represent significant obstacles for in vivo studies. To address these challenges, researchers have developed various methodologies to create model cell membranes or membrane fragments, including mono- or bilayers organized in planar systems. These models facilitate fundamental studies on membrane component interactions as well as the interactions of membrane components with external agents, such as drugs, nanoparticles (NPs), or biomarkers. The applications of model cell membranes have extended beyond basic research, encompassing areas such as biosensing and nanoparticle camouflage to evade immune detection. In this review, we highlight advancements in the engineering of planar model cell membranes, focusing on the nanoarchitectonic tools used for their fabrication. We also discuss approaches for incorporating challenging materials, such as proteins and enzymes, into these models. Finally, we present our view on future perspectives in the field of planar model cell membranes.

Simulating Bacterial Membrane Models at the Atomistic Level: A Force Field Comparison.

Molecular dynamics (MD) simulations are currently an indispensable tool to understand both the dynamic and nanoscale organization of cell membrane models. A large number of quantitative parameters can be extracted from these simulations, but their reliability is determined by the quality of the employed force field and the simulation parameters. Much of the work on parametrizing and optimizing force fields for biomembrane modeling has been focused on homogeneous bilayers with a single phospholipid type. However, these may not perform effectively or could even be unsuitable for lipid mixtures commonly employed in membrane models. This work aims to fill this gap by comparing MD simulation results of several bacterial membrane models using different force fields and simulation parameters, namely, CHARMM36, Slipids, and GROMOS-CKP. Furthermore, the hydrogen isotope exchange (HIE) method, combined with GROMOS-CKP (GROMOS-H2Q), was also tested to check for the impact of this acceleration strategy on the performance of the force field. A common set of simulation parameters was employed for all of the force fields in addition to those corresponding to the original parametrization of each of them. Furthermore, new experimental order parameter values determined from NMR of several lipid mixtures are also reported to compare them with those determined from MD simulations. Our results reveal that most of the calculated physical properties of bacterial membrane models from MD simulations are substantially force field and lipid composition dependent. Some lipid mixtures exhibit nearly ideal behaviors, while the interaction of different lipid types in other mixtures is highly synergistic. None of the employed force fields seem to be clearly superior to the other three, each having its own strengths and weaknesses. Slipids are notably effective at replicating the order parameters for all acyl chains, including those in lipid mixtures, but they offer the least accurate results for headgroup parameters. Conversely, CHARMM provides almost perfect estimates for the order parameters of the headgroups but tends to overestimate those of the lipid tails. The GROMOS parametrizations deliver reasonable order parameters for entire lipid molecules, including multicomponent bilayers, although they do not reach the accuracy of Slipids for tails or CHARMM for headgroups. Importantly, GROMOS-H2Q stands out for its computational efficiency, being at least 3 times faster than GROMOS, which is already faster than both CHARMM and Slipids. In turn, GROMOS-H2Q yields much higher compressibilities compared to all other parametrizations.

Cytotoxicity and cell membrane interactions of choline-based ionic liquids: Comparing amino acids, acetate, and geranate anions

Ionic liquids (ILs) have found diverse applications in research and industry. Biocompatible ILs, a subset considered less toxic than traditional ILs, have expanded their applications into biomedical fields. However, there is limited understanding of the toxicity profiles, safe concentrations, and underlying factors driving their toxicity. In this study, we investigated the cytotoxicity of 13 choline-based ILs using four different cell lines: Human dermal fibroblasts (HDF), epidermoid carcinoma cells (A431), cervical cancer cells (HeLa), and gastric cancer cells (AGS). Additionally, we explored the haemolytic activity of these ILs. Our findings showed that the cytotoxic and haemolytic activities of ILs can be attributed to the hydrophobicity of the anions and the pH of the IL solutions. Furthermore, utilising quartz crystal microbalance with dissipation (QCM-D), we delved into the interaction of selected ILs, including choline acetate [Cho][Ac] and choline geranate [Cho][Ge], with model cell membranes composed of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). The QCM-D data showed that ILs with higher toxicities exhibited more pronounced interactions with membranes. Increased variations in frequency and dissipation reflected substantial changes in membrane fluidity and mass following the addition of the more toxic ILs. Furthermore, total internal reflection fluorescence microscopy study revealed that [Cho][Ac] could cause lipid rearrangements and pore formation in the membrane, while [Cho][Ge] disrupted the bilayer packing. This study advances our understanding of the cellular toxicities associated with choline-based ILs and provides valuable insights into their mechanisms of action concerning IL-membrane interactions. These findings have significant implications for the safe and informed utilisation of biocompatible ILs in the realm of drug delivery and biotechnology.

Influence of Simvastatin and Pravastatin on the Biophysical Properties of Model Lipid Bilayers and Plasma Membranes of Live Cells.

Statins are among the most widely used drugs for the inhibition of cholesterol biosynthesis, prevention of cardiovascular diseases, and treatment of hypercholesterolemia. Additionally, statins also exhibit cholesterol-independent benefits in various diseases, including neuroprotective properties in Alzheimer's disease, anti-inflammatory effects in coronary artery disease, and antiproliferative activities in cancer, which likely result from the statins' interaction and alteration of lipid bilayers. However, the membrane-modulatory effects of statins and the mechanisms by which statins alter lipid bilayers remain poorly understood. In this work, we explore the membrane-modulating effects of statins on model lipid bilayers and live cells. Through the use of fluorescence lifetime imaging microscopy (FLIM) combined with viscosity-sensitive environmental probes, we demonstrate that hydrophobic, but not hydrophilic, statins are capable of changing the microviscosity and lipid order in model and live cell membranes. Furthermore, we show that hydrophobic simvastatin is capable of forming nanoscale cholesterol-rich domains and homogenizing the cholesterol concentrations in lipid bilayers. Our results provide a mechanistic framework for understanding the bimodal effects of simvastatin on the lipid order and the lateral organization of cholesterol in lipid bilayers. Finally, we demonstrate that simvastatin temporarily decreases the microviscosity of live cell plasma membranes, making them more permeable and increasing the level of intracellular chemotherapeutic drug accumulation.

Resolving the interactions between hydrophilic CdTe quantum dots and positively charged membranes at the nanoscale

The use of quantum dot nanoparticles (QDs) in bio-applications has gained quite some interest and requires a deep understanding of their interactions with model cell membranes. This involves assessing the extent of nanoparticle disruption of the membrane and how it depends on both nanoparticle and membrane physicochemical properties. Surface charge plays an important role in nanoparticle adsorption, which is primarily driven by electrostatic interactions; yet, once adsorbed, most reported works overlook the subsequent spatial nanoparticle insertion and location within the membrane. There is therefore a need for studies to assess the mutual role of membrane and nanoparticle charge into membrane structure and stability at the nanoscale, with a view to better design and control the functionality of these nanomaterials. In this work, we have resolved the extent of the interactions between hydrophilic, negatively charged CdTe QDs and positively charged lipid bilayers. A multiscale combination of surface-sensitive techniques enabled probing how surface charge mediates QD adsorption and membrane reorganization. Increasing membrane surface charge results into a larger adsorption of oppositely charged QDs, concomitantly inducing structural changes. Hydration of the membrane hydrophobic parts by QDs goes deeper into the inner leaflet with increasing membrane charge, resulting in supported lipid bilayers with decreased nanomechanical stability.

A systematic study of the impact of aromatic/aliphatic amines and protein corona as coatings of iron oxide magnetic nanoparticles on the interaction with DPPC Langmuir monolayers

The nature of chemical groups exposed by nanoparticles determines their interaction with biological systems, especially with cell membranes. Amino groups are one of the most used to guide nanoparticles within cells. Aromatic and aliphatic amines can establish different types of interactions with lipids, and this may condition the nanoparticles' mode of action as well as their fate. This work is focused on investigating how aromatic and aliphatic amines present at the surface of iron oxide magnetic nanoparticles influence their interaction with Langmuir monolayers as models of cell membranes. For this, we synthesized iron oxide magnetic nanoparticles coated with an arylamine (aromatic) and polyethylenimine (aliphatic). Also, the effect of human serum albumin at the particle surface was evaluated to test how protein corona governs the physicochemical properties of the nanoparticles. The interaction of these nanoparticles with monolayers of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine was systematically studied employing three different experimental approaches. The results signal a notorious influence on the type of amine in the interactions established with the phosphocholine in the model membrane. Managing this knowledge is of ongoing interest for nanomaterials design and improved bioapplications.

Interaction of chondroitin sulfate with zwitterionic lipid membranes

Chondroitin sulfates (CSs) are important components of the extracellular matrix and side chains of membrane proteoglycans. These polysaccharides are, therefore, likely to interact with plasma membranes and play a significant role in modulating cellular functions. So far, the details of the processes occurring at the interface between the extracellular matrix and cellular membranes are not fully understood. In this study, we used experimental methods and atomic-scale molecular dynamics (MD) simulations to reveal the molecular picture of the interactions between CS and phosphocholine (PC) membranes, used as a simplified model of cell membranes. MD simulations reveal that the polysaccharide associates to the PC bilayer as a result of electrostatic interactions between the positively charged quaternary ammonium groups of choline and the negatively charged sulfate groups of CS. Compared to an aqueous medium, the adsorbed polysaccharide chains adopt more elongated conformations, which facilitates the electrostatic interactions with the membrane, and have a high degree of freedom to change their conformations and to adhere to and detach from the membrane surface. Penetrating slightly between the polar groups of the bilayer, they form a loosely anchored layer, but do not intrude into the hydrophobic region of the PC bilayer. The CS adsorption spread the PC headgroups apart, which is manifested by an increase in the value of the area pre lipid. The expansion of the lipid polar groups weakens the dispersion interactions between the lipid acyl chains. As a result, the lipid membrane in the membrane-polysaccharide contact areas becomes more fluid. Our outcomes may help to understand in detail the interaction of chondroitin sulfate with zwitterionic membranes at the molecular level, which is of biological interest since many biological processes depend on lipid-CS interactions.

Calorimetric and Raman spectroscopic studies of zwitterionic and anionic membrane interactions of phenolic compound coumarin

It is well known that polyphenols possess potent antioxidant qualities. However, the precise antioxidant action mechanism of phenolic compounds is still not well defined. It is evident that understanding the molecular mechanisms behind polyphenol-lipid interactions is necessary to comprehend this impact. The objective of this work is to examine the effect of phenolic compound coumarin on the physicochemical properties of cell membrane models. To accomplish this goal, model membranes made up of zwitterionic dimyristoylphosphatidylcholine (DMPC) and anionic dimyristoylphosphatidylglycerol (DMPG) lipids were used. These lipid species are the main component of the mammalian and gram-positive bacteria membranes, respectively. Differential scanning calorimetry (DSC) and Raman spectroscopy were applied for studying the interaction of coumarin with DMPC and DMPG. From the DSC results, it was found that pretransition was abolished for both lipid systems at 20 mol% coumarin. Main phase transition shifted to lower temperatures and broadened, but there was not a complete disappearance of main transition and coumarin was not fully miscible. A sharp peak at a higher temperature and a broad shoulder at a lower temperature of the main transition of DMPC were detected when compared to coumarin/DMPG binary system. A reduction in calorimetric enthalpy and entropy values of coumarin/DMPC system at 20 mol% was observed whereas an increase in these calorimetric parameters for coumarin/DMPG system occurred. Thus, the disorder was caused when 20 mol% coumarin interacts with DMPC lipid bilayers. The increase in enthalpy could be a result of the 20 mol% coumarin interactions with DMPG lipid bilayers or a possible generated partial interdigitation. From the Raman results, the peak height Raman intensity ratio I1090/I1130 showed that coumarin induces disorder in the DMPG bilayers in the gel phase due to the increasing gauche:trans ratio. According to the results obtained by DSC and Raman spectroscopy, it was found that coumarin has a varied effect on membranes made from lipids with choline and glycerol head groups. Thus, it is obvious that the polyphenol/membrane interactions can be significantly impacted by the lipids’ structural variations.

Unsaturation effects on lipid transmembrane asymmetry.

Within cell plasma membranes, unsaturated lipids are asymmetrically distributed over the inner and outer leaflets, offering an attractive local structural feature. However, the mechanism to keep lipid transmembrane asymmetry and the closely related transmembrane movement (flip-flop) for unsaturated lipids remain poorly understood. Here, we applied sum frequency generation vibrational spectroscopy to investigate this lipid transmembrane asymmetry upon mimicking the cell membrane homeostatic processes. On the one hand, unsaturated lipids were found to hinder the flip-flop process and preserve lipid transmembrane asymmetry in model cell membranes, owing to the steric hindrance caused by their bent tails. On the other hand, local unsaturated lipids in the mixed unsaturated/saturated lipid bilayer were conducive to the formation of the local asymmetry. Therefore, lipid unsaturation can be recognized as an intrinsic key factor to form and maintain lipid transmembrane asymmetry in cell membranes.

Effects of inserting ultrashort carbon nanotubes into lipid bilayers on membrane morphology

AbstractSingle‐walled carbon nanotubes (CNTs) are carbon materials with unique thermal, optical, mechanical, and electrical properties, with hollow cylindrical structures of a few nanometers in diameter. CNTs cut to about 10 nm (Ultrashort CNTs, US‐CNTs) can spontaneously insert into lipid bilayers. Therefore, applications have been proposed to combine CNTs with lipid bilayers to give the membranes the properties of CNTs. However, CNTs interact with membranes to induce morphological changes in the membranes, which may hinder these applications. In this study, to investigate the effects, US‐CNTs are exposed to lipid bilayer vesicles (giant unilamellar vesicles, GUVs), which are used as a model for cell membranes, and the changes in membrane morphology with each US‐CNT concentration were evaluated by fluorescence microscopy. As a result, GUVs show morphological changes upon exposure to US‐CNTs, eventually transforming into a multiple vesicle‐linked shape. This result suggests an increase in the area and asymmetry of the GUV membrane. Based on these results, we have proposed a hypothesis regarding the mechanism of morphological changes induced in the GUV membranes by US‐CNTs exposure.

Membrane localization of actin filaments stabilizes giant unilamellar vesicles against external deforming forces

Actin organization is crucial for establishing cell polarity, which influences processes such as directed cell motility and division. Despite its critical role in living organisms, achieving similar polarity in synthetic cells remains challenging. In this study, we employ a bottom-up approach to investigate how molecular crowders facilitate the formation of cortex-like actin networks and how these networks localize and organize based on membrane shape. Using giant unilamellar vesicles (GUVs) as models for cell membranes, we show that actin filaments can arrange along the membrane to form cortex-like structures. Notably, this organization is achieved using only actin and crowders as a minimal set of components. We utilize surface micropatterning to examine actin filament organization in deformed GUVs adhered to various pattern shapes. Our findings indicate that at the periphery of spherical GUVs, actin bundles align along the membrane. However, in highly curved regions of adhered GUVs, actin bundles avoid crossing the highly curved edges perpendicular to the adhesion site and instead remain in the lower curved regions by aligning parallel to the micropatterned surface. Furthermore, the actin bundles increase the stiffness of the GUVs, effectively counteracting strong deformations when GUVs adhere to micropatterns. This finding is corroborated by real-time deformability cytometry on GUVs with synthetic actin cortices. By precisely manipulating the shape of GUVs, our study provides a minimal system to investigate the interplay between actin structures and the membrane. Our findings provide insights into the spatial organization of actin structures within crowded environments, specifically inside GUVs that resemble the size and shape of cells. This study advances our understanding of actin network organization and functionality within cell-sized compartments.

Формирование монослоев Ленгмюра из нативных фосфолипидов бактерий различных систематических групп

Phospholipids are the most important structural elements of the bacterial cell wall, participate in the adaptation of microorganisms to the environment and can act as biomarkers for environmental changes and one of the components of environmental monitoring. Native phospholipids are used to form models of cell membranes, the biophysical properties of which can be studied by the Langmuir-Blodgett method. The aim of this work was to isolate and characterize the phospholipids of the cell membranes of the bacteria Staphylococcus aureus and Bacillus cereus and the formation of Langmuir monolayers based on them. The composition and ratio of fatty acids were determined by gas-liquid chromatography of fatty acid methyl esters. Fatty acids, found in the extract of the bacteria S. aureus 209-P and B. cereus 8035, are: hexadecanoic, trans-9-octadecenoic, octadecanoic, tetradecanoic, 13-methyltetradecanoic, 14-methylpentadecanoic, 15-methylhexadecanoic, cis-9-octadecanoic. To form a monolayer, a working solution of native phospholipids in chloroform with a concentration of C = 10-3 М was used. The monolayer formed when a 50 μl solution of a phospholipid mixture is applied to the surface has a more perfect structure, which is manifested in the constancy of its mechanical properties. The analysis of the obtained data has not yet revealed a clear dependence of the monolayer parameters on temperature. The changes in the compression modulus and compressibility were very minor. With an increase in the salt concentration, both an increase and a decrease in the compression modulus, and, consequently, the rigidity of the monolayer, is observed.

Cryptotanshinone-Induced Permeabilization of Model Phospholipid Membranes: A Biophysical Study.

The Danshen terpenoid cryptotanshinone (CPT) is gaining enormous interest in light of its various outstanding biological activities. Among those, CPT has been shown to interact with cell membranes and, for instance, to have antibacterial activity. Several works have shown that CPT alone, or in combination with other drugs, can effectively act as an antibiotic against various infectious bacteria. Some authors have related the mechanism underlying this action to CPT-membrane interaction. This work shows that CPT readily partitions into phosphatidylcholine membranes, but there is a limiting capacity of accommodation of ca. 1 mol CPT to 3 mol phospholipid. The addition of CPT to unilamellar liposomes composed of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) causes membrane permeabilization, as shown by fluorescent probe leakage. This process has been kinetically studied, as well as its modulation by incorporation of phosphatidylethanolamine or phosphatidylglycerol, as a model for pathogenic cell membranes. The thermotropic behavior of 1,2-dimyristoylphosphatidylcholine (DMPC) model membranes is weakly affected by CPT, but the terpenoid causes significant dehydration of the polar region of the bilayer and weak disordering of the acyl chain palisade, as observed in Fourier-transform infrared spectroscopy (FTIR) results. Small-angle X-ray scattering (SAXS) shows that CPT increases DMPC bilayer thickness, which could be due to localization near the phospholipid/water interface. Molecular dynamics (MD) simulations show that the lateral diffusion coefficient of the phospholipid increases with the presence of CPT. CPT extends from the polar head region to the center of the bilayer, being centered between the carbonyl groups and the unsaturated region of the POPC, where there is greater overlap. Interestingly, the free energy profiles of a water molecule crossing the lipid membrane show that the POPC membrane becomes more permeable in the presence of CPT. In summary, our results show that CPT perturbs the physicochemical properties of the phospholipid membrane and compromises its barrier function, which could be of relevance to explain part of its antimicrobial or anticancer activities.

Size- and Shape-Dependent Interactions of Lipid-Coated Silver Nanoparticles: An Improved Mechanistic Understanding through Model Cell Membranes and In Vivo Toxicity.

The widespread use of silver nanoparticles (AgNPs) in various applications and industries has brought to light the need for understanding the complex relationship between the physicochemical properties (shape, size, charge, and surface chemistry) of AgNPs that affect their ability to enter cells and cause toxicity. To evaluate their toxicological outcomes, this study systematically analyzed a series of homogeneous hybrid lipid-coated AgNPs spanning sizes from 5 to 100 nm with diverse shapes (spheres, triangles, and cubes). The hybrid lipid membrane comprises hydrogenated phosphatidylcholine (HPC), sodium oleate (SOA), and hexanethiol (HT), which shield the AgNP surface from surface oxidation and toxic Ag+ ion release to minimize its contribution to toxicity. To reduce any significant effects by surface chemistry, the HPC, SOA, and HT membrane composition ratio was kept constant, and the AgNPs were assessed using embryonic zebrafish (Danio rerio). While a direct comparison cannot be drawn due to the lack of complementary sizes below 40 nm for triangular plates and cubes due to synthetic challenges, significant mortality was observed for spherical AgNPs (AgNSs) of 5, 20, 40, and 60 nm at 120 h postfertilization at concentrations ≥6 mg Ag/L. In contrast, the 10, 80, and 100 nm AgNSs, 40, 70, and 100 nm triangular plate AgNPs (AgNPLs), and 55, 75, and 100 nm cubic AgNPs (AgNCs) showed no significant mortality at 5 days postfertilization following exposure to AgNPs at concentrations up to 12 mg Ag/L. With constant surface chemistry on the AgNPs, size is the dominant factor driving toxicological responses, with smaller nanoparticles (5 to 60 nm) being the most toxic. Larger AgNSs, AgNCs, and AgNPLs from 75 to 100 nm do not show any evidence of toxicity. However, when closely examining sizes between 40 and 60 nm for AgNSs, AgNCs, and AgNPLs, there is evidence that discriminates shape as a driver of toxicity since sublethal responses generally were observed to follow a pattern, suggesting toxicity is most significant for AgNSs followed by AgNPLs and then AgNCs, which is the least toxic. Sum frequency generation vibrational spectroscopy showed that irrespective of size or shape, all hybrid lipid-coated AgNPs interact with membrane surfaces and "snorkel" between phases into the lipid monolayer with minimal energetic cost. These findings decisively demonstrate that not only smaller AgNPs but also the shape of the AgNPs influences their biological compatibility.