Search Results (31)

This mini-review intends to highlight the importance of bilayer asymmetry. Biological membranes are complex structures that are a physical barrier separating the external environment from the cellular content. This complex bilayer comprises an extensive lipid repertory, suggesting that the different lipid structures might play a role in the membrane. Interestingly, this vast repertory of lipids is asymmetrically distributed between leaflets that form the lipid bilayer. Here, we discuss the properties of the plasma membrane from the perspective of experimental model membranes, consisting of simplified and controlled in vitro systems. We summarize some crucial features of the exoplasmic (outer) and cytoplasmic (inner) leaflets observed through investigations using symmetric and asymmetric membranes. Symmetric model membranes for the exoplasmic leaflet have a unique lipid composition that might form a coexistence of phases, namely the liquid disordered and liquid order phases. These phase domains may appear in different sizes and shapes depending on lipid composition and lipid–lipid interactions. In contrast, symmetric model membranes for the cytoplasmic leaflet form a fluid phase. We discuss the outcomes reported in the literature for asymmetric bilayers, which vary according to lipid compositions and, consequently, reflect different intra- and inter-leaflet interactions. Interestingly, the asymmetric bilayer could show induced domains in the inner leaflet, or it could decrease the tendency of the outer leaflet to phase separation. If cells regulate the lipid composition of the plasma membrane, they can adjust the existence and sizes of the domains by tuning the lipid composition. Full article

Mixtures of two phospholipids (PLs) with different main phase transition temperatures were investigated. Host PLs (HPLs) were represented by DMPC, DPPC, DSPC, and DMPE. The admixed PL was the spin-labeled phosphatidylcholine 5-PC(1-palmitoyl-2-(5-doxylstearoyl)phosphatidylcholine), with a unique opportunity to monitor the properties and the local environments of all admixed PL molecules using saturation recovery EPR methods. Below the HPL phase transition temperatures, 5-PC mixes with HPL to form two distinct pools with different rotational diffusion rates. The fluidity of the local environment in these two pools is very different, being more fluid for molecules with greater rotational diffusion rates. Above the HPL phase transition temperature, 5-PC mixes with HPL uniformly. This is independent of the HPL, observed for 5-PC concentrations from 0.25 mol% up to 20 mol% and for the wide temperature range. Assuminga very low concentration of 5-PC is an ideal probe molecule, we can conclude that small fluid phase domains made of HPL molecules are formed below the phase transition temperature of the HPL bilayers. In binary mixtures of HPLs with 5-PC, below the phase transition of HPL bilayers, fluid phase domains are created within the bulk gel phase of HPL lipids by the admixed second PL, namely 5-PC. Full article

The clinical benefits of using exogenous pulmonary surfactant (EPS) as a carrier of budesonide (BUD), a non-halogenated corticosteroid with a broad anti-inflammatory effect, have been established. Using various experimental techniques (differential scanning calorimetry DSC, small- and wide- angle X-ray scattering SAXS/WAXS, small- angle neutron scattering SANS, fluorescence spectroscopy, dynamic light scattering DLS, and zeta potential), we investigated the effect of BUD on the thermodynamics and structure of the clinically used EPS, Curosurf^®. We show that BUD facilitates the Curosurf^® phase transition from the gel to the fluid state, resulting in a decrease in the temperature of the main phase transition (Tm) and enthalpy (ΔH). The morphology of the Curosurf^® dispersion is maintained for BUD < 10 wt% of the Curosurf^® mass; BUD slightly increases the repeat distance d of the fluid lamellar phase in multilamellar vesicles (MLVs) resulting from the thickening of the lipid bilayer. The bilayer thickening (~0.23 nm) was derived from SANS data. The presence of ~2 mmol/L of Ca²⁺ maintains the effect and structure of the MLVs. The changes in the lateral pressure of the Curosurf^® bilayer revealed that the intercalated BUD between the acyl chains of the surfactant’s lipid molecules resides deeper in the hydrophobic region when its content exceeds ~6 wt%. Our studies support the concept of a combined therapy utilising budesonide—enriched Curosurf^®. Full article

The dense packing of opposite cytoplasmic surfaces of the lipid-enriched myelin membrane, responsible for the proper saltatory conduction of nerve impulses through axons, is ensured by the adhesive properties of myelin basic protein (MBP). Although preferentially interacting with negatively charged phosphatidylserine (PS) lipids, as an intrinsically disordered protein, it can easily adapt its shape to its immediate environment and thus adsorb to domains made of zwitterionic phosphatidylcholine (PC) lipids. As the molecular-level interaction pattern between MBP and PC lipid membranes suffers from scarce characterization, an experimental and computational study of multilamellar liposomes (MLVs) composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) in the presence of bovine MBP is presented here. Calorimetric and temperature-dependent UV-Vis measurements identified DPPC pretransition temperature (T_p) and calorimetric enthalpy (ΔH_cal) as the physicochemical parameters most responsive to the presence of MBP. Besides suggesting an increase in β-sheet fractions of structured MBP segments as DPPC lipids undergo from the gel (20 °C) to the fluid (50 °C) phase, FTIR spectra unraveled the significant contribution of lysine (Lys) residues in the adsorption pattern, especially when DPPC is in the fluid (50 °C) phase. In addition to highlighting the importance of Lys residues in the MBP adsorption on DPPC lipid bilayer, employing salt bridges (SBs) and hydrogen bonds (HBs), MD data suggest the crucial importance of the orientation of MBP with respect to the surface of the DPPC lipid bilayer. Full article

Cell-penetrating peptides (CPPs) are short peptides built up from dominantly cationic and hydrophobic amino acid residues with a distinguished ability to pass through the cell membrane. Due to the possibility of linking and delivering the appropriate cargo at the desired location, CPPs are considered an economic and less invasive alternative to antibiotics. Besides knowing that their membrane passage mechanism is a complex function of CPP chemical composition, the ionic strength of the solution, and the membrane composition, all other details on how they penetrate cell membranes are rather vague. The aim of this study is to elucidate the ad(de)sorption of arginine-/lysine- and phenylalanine-rich peptides on a lipid membrane composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipids. DSC and temperature-dependent UV-Vis measurements confirmed the impact of the adsorbed peptides on thermotropic properties of DPPC, but in an inconclusive way. On the other hand, FTIR spectra acquired at 30 °C and 50 °C (when DPPC lipids are found in the gel and fluid phase, respectively) unambiguously confirmed the proton transfer between particular titratable functional groups of R5F2/K5F2 that highly depend on their immediate surroundings (DPPC or a phosphate buffer). Molecular dynamic simulations showed that both peptides may adsorb onto the bilayer, but K5F2 desorbs more easily and favors the solvent, while R5F2 remains attached. The results obtained in this work highlight the importance of proton transfer in the design of CPPs with their desired cargo, as its charge and composition dictates the possibility of entering the cell. Full article

Despite the undisputable role of the protein corona in the biointeractions of liposome drug carriers, the field suffers from a lack of knowledge regarding the patterns of protein deposition on lipid surfaces with different compositions. Here, we investigated the protein coronas formed on liposomes of basic compositions containing combinations of egg phosphatidylcholine (PC), palmitoyloleoyl phosphatidylglycerol (POPG), and cholesterol. Liposome−protein complexes isolated by size-exclusion chromatography were delipidated and analyzed using label-free LC-MS/MS. The addition of the anionic lipid and cholesterol both affected the relative protein abundances (and not the total bound proteins) in the coronas. Highly anionic liposomes, namely those containing 40% POPG, carried corona enriched with cationic proteins (apolipoprotein C1, beta-2-glycoprotein 1, and cathelicidins) and were the least stable in the calcein release assay. Cholesterol improved the liposome stability in the plasma. However, the differences in the corona compositions had little effect on the liposome uptake by endothelial (EA.hy926) and phagocytic cells in the culture (U937) or ex vivo (blood-derived monocytes and neutrophils). The findings emphasize that the effect of protein corona on the performance of the liposomes as drug carriers occurs through compromising particle stability rather than interfering with cellular uptake. Full article

Biological symmetry breaking is a mechanism in biosystems that is necessary for human survival, and depends on chemical physics concepts at both microscopic and macroscopic scales. In this work, we present a few mechanisms of the signaling phenomenon that have been studied in various tissues of human origin. We exhibit that anatomical asymmetry in the structure of a membrane can produce a flow of extracellular fluid. Furthermore, we exhibit that membrane asymmetry is a misbalance in the composition of the aqueous phases and interaction forces with the protein trans-membrane. Various biological membranes such as DPPC, DMPC, DLPC, and so on, have considerable electrostatic voltages that extend across the phosphor lipids bilayer. For studying these phenomena, we modeled DPPC, DMPC, and DLPC lipid bilayers with a net charge misbalance across the phospholipids. Because asymmetric membranes create the shifted voltages among the various aqueous tissues, this effect makes the charge misbalances cause a voltage of 1.3 V across the DPPC bilayer and 0.8 V across the DMPC bilayer. This subject exhibits the importance of membrane structures on electrostatic potential gradients. Finally, we exhibited that a quantum effect was created in small parts of the cell’s thickness due to the symmetry breaking of asymmetrical phospholipid bilayers. Full article

The gel-phase domains in a binary supported lipid bilayer (SLB) comprising dioleoylphosphatidylcholine (DOPC) and dipalmitoylphosphatidylcholine (DPPC) were localized on graphene oxide (GO) deposited on a SiO₂/Si substrate. We investigated the distribution of the gel-phase domains and the liquid crystalline (L_α) phase regions in DOPC+DPPC-SLB on thermally oxidized SiO₂/Si substrates with GO flakes to understand the mechanism of the domain localization on GO. Fluorescence microscopy and atomic force microscopy revealed that the gel-phase domains preferably distributed on GO flakes, whereas the fraction of the L_α-phase increased on the bare SiO₂ surface which was not covered with the GO flakes. The gel-phase domain was condensed on GO more effectively at the lower cooling rate. We propose that nucleation of the gel-phase domain preferentially occurred on GO, whose surface has amphiphilic property, during the gel-phase domain formation. The domains of the liquid ordered (L_o) phase were also condensed on GO in a ternary bilayer containing cholesterol that was phase-separated to the L_o phase and the liquid disordered phase. Rigid domains segregates on GO during their formation process, leaving fluid components to the surrounding region of GO. Full article

In this study, the interfacial behavior of high internal phase emulsions (HIPEs), stabilized by ultrasound combined with pH-shifting modified pea protein isolate (MPPI), was investigated, and its emulsification process and stabilization mechanism were discussed. The effects of MPPI concentration on the micromorphology, droplet size, rheology, and stability of HIPEs were investigated. As the MPPI concentration increased, the appearance of HIPEs gradually changed from a relatively fluid state to a plastic solid-like state with detailed texture. There occurred a gradual decrease in droplet size, the cohering of an orderly and tight arrangement, in addition to the formation of a bilayer elastic interface layer. The macro- and microrheological assessments confirmed that the apparent viscosity, storage modulus, elasticity index, and macroscopic viscosity index increased gradually. Furthermore, it was demonstrated that 5 wt% MPPI-stabilized HIPEs had the potential to be used as 3D printing inks. Stability evaluation showed that the TURBISCAN stability index decreased and centrifugal stability increased. The appearance and microstructure remained highly stable after heating at 80 °C for 30 min and storage at 4 ℃ for 90 days. These findings confirm that MPPI improves the rheological behavior and stability of HIPEs by modulating the interfacial adsorption and network structure. Full article

Cardiolipin (CL) is a key lipid for damaged mitochondrial recognition by the LC3/GABARAP human autophagy proteins. The role of ceramide (Cer) in this process is unclear, but CL and Cer have been proposed to coexist in mitochondria under certain conditions. Varela et al. showed that in model membranes composed of egg sphingomyelin (eSM), dioleoyl phosphatidylethanolamine (DOPE), and CL, the addition of Cer enhanced the binding of LC3/GABARAP proteins to bilayers. Cer gave rise to lateral phase separation of Cer-rich rigid domains but protein binding took place mainly in the fluid continuous phase. In the present study, a biophysical analysis of bilayers composed of eSM, DOPE, CL, and/or Cer was attempted to understand the relevance of this lipid coexistence. Bilayers were studied by differential scanning calorimetry, confocal fluorescence microscopy, and atomic force microscopy. Upon the addition of CL and Cer, one continuous phase and two segregated ones were formed. In bilayers with egg phosphatidylcholine instead of eSM, in which the binding of LC3/GABARAP proteins hardly increased with Cer in the former study, a single segregated phase was formed. Assuming that phase separation at the nanoscale is ruled by the same principles acting at the micrometer scale, it is proposed that Cer-enriched rigid nanodomains, stabilized by eSM:Cer interactions formed within the DOPE- and CL-enriched fluid phase, result in structural defects at the rigid/fluid nanointerfaces, thus hypothetically facilitatingLC3/GABARAP protein interaction. Full article

Cyt proteins are insecticidal proteins originally from Bacillus thuringiensis. The lipid binding of the Cyt2Aa2 protein depends on the phase of the lipid bilayer. In this work, the importance of the conserved T144 residue in the αD-β4 loop for lipid binding on fluid lipid membranes was investigated via atomic force microscopy (AFM). Lipid membrane fluidity could be monitored for the following lipid mixture systems: POPC/DPPC, POPC/SM, and DOPC/SM. AFM results revealed that the T144A mutant was unable to bind to pure POPC bilayers. Similar topography between the wildtype and T144A mutant was seen for the POPC/Chol system. Small aggregates of T144A mutant were observed in the POPC and DOPC domains of the lipid mixture systems. In addition, the T144A mutant had no cytotoxic effect against human colon cancer cells. These results suggest that alanine replacement into threonine 144 hinders the binding of Cyt2Aa2 on liquid lipid membranes. These observations provide a possibility to modify the Cyt2Aa2 protein to specific cells via lipid phase selection. Full article

Lipids that have two tails of different lengths are found throughout biomembranes in nature, yet the effects of this asymmetry on the membrane properties are not well understood, especially when it comes to the membrane dynamics. Here we study the nanoscale bending fluctuations in model mixed-chain 14:0–18:0 PC (MSPC) and 18:0–14:0 PC (SMPC) lipid bilayers using neutron spin echo (NSE) spectroscopy. We find that despite the partial interdigitation that is known to persist in the fluid phase of these membranes, the collective fluctuations are enhanced on timescales of tens of nanoseconds, and the chain-asymmetric lipid bilayers are softer than an analogous chain-symmetric lipid bilayer with the same average number of carbons in the acyl tails, di-16:0 PC (DPPC). Quantitative comparison of the NSE results suggests that the enhanced bending fluctuations at the nanosecond timescales are consistent with experimental and computational studies that showed the compressibility moduli of chain-asymmetric lipid membranes are 20% to 40% lower than chain-symmetric lipid membranes. These studies add to growing evidence that the partial interdigitation in mixed-chain lipid membranes is highly dynamic in the fluid phase and impacts membrane dynamic processes from the molecular to mesoscopic length scales without significantly changing the bilayer thickness or area per lipid. Full article

Bile acids form micelles that are essential for the absorption of dietary lipids. However, excessive bile acid micelles can disrupt the plasma membrane by removing phospholipids, resulting in cell death. We hypothesized that the bent geometrical structure of the steroid scaffold of bile acids decreases the lipid order (similar to unsaturated phospholipids with cis double bonds), disrupting the plasma membrane. Here, lithocholic acid (LCA), a bile acid, was methylated to prevent micellization. Methylated lithocholic acid (Me-LCA) was mixed with a thin phase-separated lipid bilayer comprising 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), and cholesterol (Chol). Me-LCA was not localized in the DPPC-rich rigid phase but localized in the DOPC-rich fluid phase, and excess Me-LCA did not affect the phase separation. Me-LCA is distributed in the plasma and organelle membranes. However, Me-LCA with bent structure did not affect the membrane properties, membrane fluidity, and hydrophobicity of liposomes composed of DOPC, DPPC, and Chol and also did not affect the proliferation of cells. Full article

In recent years, significant progress has been made in the development of new technologies to meet the demand for engineered interfaces with appropriate properties for osteochondral unit repair and regeneration. In this context, we combined two methodologies that have emerged as powerful approaches for tissue engineering application: electrospinning to fabricate a nanofibrous polymeric scaffold and pulsed laser deposition to tune and control the composition and morphology of the scaffold surface. A multi-component scaffold composed of synthetic and natural polymers was proposed to combine the biocompatibility and suitable mechanical properties of poly(D,L-lactic acid) with the hydrophilicity and cellular affinity of gelatin. As part of a biomimetic strategy for the generation of bi-functional scaffolds, we coated the electrospun fibers with a thin film of a bioactive glass–ceramic material supplemented with manganese ions. The physico-chemical properties and composition of the bi-layered scaffold were investigated, and its bioactivity, in terms of induced mineralization, was tested by incubation in a simulated body fluid buffer. The processes of the inorganic film dissolution and the calcium phosphate phases growth were followed by microscopic and spectroscopic techniques, confirming that a combination of bioactive glass–ceramics and nanofibrous scaffolds has promising potential in the regeneration of osteochondral tissue due to its ability to induce mineralization in connective tissues. Full article

In the last years, a few experiments in the fields of biological and soft matter physics in colloidal suspensions have reported “normal diffusion” with a Laplacian probability distribution in the particle’s displacements (i.e., Brownian yet non-Gaussian diffusion). To model this behavior, different stochastic and microscopic models have been proposed, with the former introducing new random elements that incorporate our lack of information about the media and the latter describing a limited number of interesting physical scenarios. This incentivizes the search of a more thorough understanding of how the media interacts with itself and with the particle being diffused in Brownian yet non-Gaussian diffusion. For this reason, a comprehensive mathematical model to explain Brownian yet non-Gaussian diffusion that includes weak molecular interactions is proposed in this paper. Based on the theory of interfaces by De Gennes and Langevin dynamics, it is shown that long-range interactions in a weakly interacting fluid at shorter time scales leads to a Laplacian probability distribution in the radial particle’s displacements. Further, it is shown that a phase separation can explain a high diffusivity and causes this Laplacian distribution to evolve towards a Gaussian via a transition probability in the interval of time as it was observed in experiments. To verify these model predictions, the experimental data of the Brownian motion of colloidal beads on phospholipid bilayer by Wang et al. are used and compared with the results of the theory. This comparison suggests that the proposed model is able to explain qualitatively and quantitatively the Brownian yet non-Gaussian diffusion. Full article