Lipid Phase Research Articles

Mechanisms underlying lipid emulsion resuscitation for drug toxicity: a narrative review.

Currently, lipid emulsion (LE) is widely used in the treatment of local anesthetic systemic toxicity (LAST). Additionally, LE has been shown to be effective to ameliorate intractable cardiovascular collapse evoked by toxicity of lipid soluble non-local anesthetic drug. The goal of this review focused on the underlying mechanism of LE treatment in the drug toxicity including LAST, LE treatment, and further research direction. We searched the relevant articles using following keywords: "local anesthetic systemic toxicity or LAST or toxicity or intoxication or poisoning" and "Intralipid or lipid emulsion". The underlying mechanism associated with LE treatment can be divided into indirect and direct effects. Lipid shuttle (indirect effect) of LE treatment is the commonly accepted mechanism: lipid phase of LE absorbs highly lipid soluble drugs ( bupivacaine) from heart and brain, and then LE containing lipid soluble drugs is delivered to muscle, adipose tissue, and liver for storage and detoxification. The direct effects are as follows: inotropic effects, supply of fatty acid, attenuation of mitochondrial dysfunction, glycogen synthase kinase-3β phosphorylation, and inhibition of nitric oxide. These mechanisms seem to synergistically act to alleviate drug toxicity. The recommended LE dosage for LAST is as follows: bolus administration of 20% LE 1.5 ml/kg over 2 to 3 min followed by 20% LE infusion 0.25 ml/kg/min. LAST most occurs via intravenous administration. However, as drug toxicity caused by non-local anesthetic drugs occurs via oral administration, further study is needed to examine the optimal dosing schedule of LE treatment for non-local anesthetic drug toxicity.

Neuroprotective Effect of α-Lipoic Acid against Aβ25-35-Induced Damage in BV2 Cells.

The prevalence of Alzheimer's disease (AD) is significantly increasing due to the aging world population, and the currently available drug treatments cannot cure or even slow its progression. α-lipoic acid (LA) is a biological factor widely found in spinach and meat and can dissolve in both lipid and aqueous phases. In medicine, LA has been shown to reduce the symptoms of diabetic polyneuropathy, acute kidney injury, cancers, and some metabolism-related diseases. This study to proves that α-lipoic acid (LA) can stabilize the cognitive function of patients with Alzheimer's disease (AD). BV2 cells were divided into control, LA, Aβ25-35, and LA + Aβ25-35 groups. Cell growth; IL-6, IL-1β, TNF-α, IFN-γ, SOD, GPx, CAT, ROS, NO, and iNOS secretion; Wnt-related proteins; cell apoptosis; and cell activation were examined. Here, we found that LA could effectively repress apoptosis and changes in the morphology of microglia BV2 cells activated by Aβ25-35, accompanied by the inhibition of the inflammatory response induced by Aβ25-35. The Wnt/β-catenin pathway is also involved in preventing Aβ25-35-induced cytotoxicity in microglia by LA. We found an inhibitory effect of LA on microglia toxicity induced by Aβ25-35, suggesting that a combination of anti-inflammatory and antioxidant substances may offer a promising approach to the treatment of AD.

Theoretical and Experimental Study of Molecular Interactions of Fluralaner with Lipid Membranes.

Fluralaner is a relatively new insecticide belonging to the isoxazoline group, whose action mechanism involves the blocking of GABAA-receptors in the insect nervous system. Because of its high hydrophobicity, fluralaner could bioaccumulate and reach toxic local concentrations. Since there are no data available about the penetration and persistence of isoxazolines in biological membranes, we intend to evaluate fluralaner permanence as a pollutant by using model membranes. We used experimental and in silico models to characterize the incorporation of fluralaner into the lipid phase at different packing states. We determined its impact in the membrane structure and organization. Our results confirm that fluralaner is capable of penetrating, holding, and accumulating in the lipid membrane and provide details on its precise location and orientation. These properties would allow fluralaner to reach high local concentrations in different membranes and organs, which could be dangerous for vertebrate organisms if its handling is not properly controlled.

Interactions between DMPC Model Membranes, the Drug Naproxen, and the Saponin β-Aescin.

In this study, the interplay among the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) as a model membrane, the nonsteroidal anti-inflammatory drug naproxen, and the saponin β-aescin are investigated. The naproxen amount was fixed to 10 mol%, and the saponin amount varies from 0.0 to 1.0 mol%. Both substances are common ingredients in pharmaceutics; therefore, it is important to obtain deeper knowledge of their impact on lipid membranes. The size and properties of the DMPC model membrane upon naproxen and aescin addition were characterized with differential scanning calorimetry (DSC), small- and wide-angle X-ray scattering (SAXS, WAXS), and photon correlation spectroscopy (PCS) in a temperature-dependent study. The interaction of all substances was dependent on the lipid phase state, which itself depends on the lipid's main phase transition temperature Tm. The incorporation of naproxen and aescin distorted the lipid membrane structure and lowers Tm. Below Tm, the DMPC-naproxen-aescin mixtures showed a vesicle structure, and the insertion of naproxen and aescin influenced neither the lipid chain-chain correlation distance nor the membrane thickness. Above Tm, the insertion of both molecules instead induced the formation of correlated bilayers and a decrease in the chain-chain correlation distance. The presented data clearly confirm the interaction of naproxen and aescin with DMPC model membranes. Moreover, the incorporation of both additives into the model membranes is evidenced.

New food, new technology: innovative spreadable cream with strawberry syrup

A strawberry spreadable cream was developed, valorizing regional raw materials, contributing to food waste reduction and agri-food ecosystem sustainability. Spreadable creams are water-in-oil emulsions whose lipid phase normally contains a blend of vegetable oils, natural colourants, stabilizers, emulsifiers, flavourings, antioxidants, lecithin, and fat-soluble vitamins. The aqueous phase normally contains skim milk proteins and small quantities of other ingredients, such as salt, preservatives, thickeners, and water-soluble vitamins. The methodology involved the experimental technological development articulated with microbiological, proximal, physicochemical, and sensorial analysis. This new product revealed nutritional advantages over similar products already on the market. The final prototype was subjected to food pairing and food design with incremental acceptance according to gastronomic use, in addition to its direct use as a spreadable cream. This work was part of the project Agrio et Emulsio—new products development (POCI-01-0145-FEDER-023583), whose main objective was the formulation and design of innovative food emulsions based on processed raw materials, with potential application in certain markets such as gourmet, diet, and vegan.

Inulin-lipid hybrid (ILH) microparticles promote pH-triggered release of rifampicin within infected macrophages

Intracellular bacteria serve as a problematic source of infection due to their ability to evade biological immune responses and the inability for conventional antibiotics to efficiently penetrate cellular membranes. Subsequently, new treatment approaches are urgently required to effectively eradicate intracellular pathogens residing within immune cells (e.g. macrophages). In this study, the poorly soluble and poorly permeable antibiotic, rifampicin, was re-purposed via micro-encapsulation within inulin-lipid hybrid (ILH) particles for the treatment of macrophages infected with small colony variants of Staphylococcus aureus (SCV S. aureus). Rifampicin-encapsulated ILH (Rif-ILH) microparticles were synthesized by spray drying a lipid nano-emulsion, with inulin dissolved throughout the aqueous phase and rifampicin pre-loaded within the lipid phase. Rif-ILH were strategically designed and engineered with pH-responsive properties to promote lysosomal drug release upon cellular internalization, while preventing premature rifampicin release in plasma-simulating media. The pH-responsiveness of Rif-ILH was controlled by the acid-mediated hydrolysis of the inulin coating, where exposure to acidic media simulating the lysosomal environment of macrophages triggered hydrolysis of the oligofructose chain and the subsequent diffusion of rifampicin from Rif-ILH. This pH-provoked release mechanism, as well as the ability for ILH microparticles to be more readily internalized by macrophages, was found to be influential in triggering a 2.9-fold increase in intracellular rifampicin concentration within infected macrophages, compared to the pure drug. The subsequent increase in exposure of intracellular pathogens to rifampicin leads to a ~ 2-log improvement in antibacterial activity for Rif-ILH, at a rifampicin dose of 2.5 µg/mL. Thus, the reduction in viability of intracellular SCV S. aureus, in the absence of cellular toxicity, is indicative of ILH microparticles serving as a unique approach for the safe and efficacious delivery of antibiotics to phagocytic cells for the treatment of intracellular infections.Graphical

Temperature-Dependent Re-alignment of the Short Multifunctional Peptide BP100 in Membranes Revealed by Solid-State NMR Spectroscopy and Molecular Dynamics Simulations.

BP100 is a cationic undecamer peptide with antimicrobial and cell-penetrating activities. The orientation of this amphiphilic α-helix in lipid bilayers was examined under numerous conditions using solid-state 19 F, 15 N and 2 H NMR. At high temperatures in saturated phosphatidylcholine lipids, BP100 lies flat on the membrane surface, as expected. Upon lowering the temperature towards the lipid phase transition, the helix is found to flip into an upright transmembrane orientation. In thin bilayers, this inserted state was stable at low peptide concentration, but thicker membranes required higher peptide concentrations. In the presence of lysolipids, the inserted state prevailed even at high temperature. Molecular dynamics simulations suggest that BP100 monomer insertion can be stabilized by snorkeling lysine side chains. These results demonstrate that even a very short helix like BP100 can span (and thereby penetrate through) a cellular membrane under suitable conditions.

Nanoscale membrane curvature sorts lipid phases and alters lipid diffusion

The precise spatiotemporal control of nanoscale membrane shape and composition is the result of a complex interplay of individual and collective molecular behaviors. Here, we employed single-molecule localization microscopy and computational simulations to observe single-lipid diffusion and sorting in model membranes with varying compositions, phases, temperatures, and curvatures. Supported lipid bilayers were created over 50-nm-radius nanoparticles to mimic the size of naturally occurring membrane buds, such as endocytic pits and the formation of viral envelopes. The curved membranes recruited liquid-disordered lipid phases while altering the diffusion and sorting of tracer lipids. Disorder-preferring fluorescent lipids sorted to and experienced faster diffusion on the nanoscale curvature only when embedded in a membrane capable of sustaining lipid phase separation at low temperatures. The curvature-induced sorting and faster diffusion even occurred when the sample temperature was above the miscibility temperature of the planar membrane, implying that the nanoscale curvature could induce phase separation in otherwise homogeneous membranes. Further confirmation and understanding of these results are provided by continuum and coarse-grained molecular dynamics simulations with explicit and spontaneous curvature-phase coupling, respectively. The curvature-induced membrane compositional heterogeneity and altered dynamics were achieved only with a coupling of the curvature with a lipid phase separation. These cross-validating results demonstrate the complex interplay of lipid phases, molecular diffusion, and nanoscale membrane curvature that are critical for membrane functionality.

Chemical composition, antimicrobial and antioxidant activity of a "lipid phase" from zanthoxylum pistaciifolium griseb leaves

Chemical composition, antimicrobial and antioxidant activity of a "lipid phase" from zanthoxylum pistaciifolium griseb leaves

Optimization of hyaluronan-enriched cubosomes for bromfenac delivery enhancing corneal permeation: characterization, ex vivo, and in vivo evaluation

To design and evaluate hyaluronan-based cubosomes loaded with bromfenac sodium (BS) for ocular application to enhance the corneal permeation and retention in pterygium and cataract treatment. BS-loaded cubosomes were prepared by the emulsification method, employing 23 full factorial design using Design-Expert® software. Glycerol monoolein (GMO) and poloxamer 407 (P407) as lipid phase and polyvinyl alcohol (PVA) as stabilizer were the used ingredients. The optimized formulation (OBC; containing GMO (7% w/w), P407 (0.7% w/w) and PVA (2.5% w/w)) was further evaluated. OBC had an entrapment efficiency of 61.66 ± 1.01%, a zeta potential of −30.80 ± 0.61 mV, a mean particle size of 149.30 ± 15.24 nm and a polydispersity index of 0.21 ± 0.02. Transmission electron microscopy confirmed its cubic shape and excellent dispersibility. OBC exhibited high stability and no ocular irritation that was ensured by histopathology. Ex vivo permeation study showed a significant increase in drug deposition and permeability parameters through goat cornea, besides, confocal laser microscopy established the superior permeation capability of OBC, as compared to drug solution. In vivo pharmacokinetics in aqueous humor indicated higher AUC0-tlast (18.88 µg.h/mL) and mean residence time (3.16 h) of OBC when compared to the marketed eye drops (7.93 µg.h/mL and 1.97 h, respectively). Accordingly, hyaluronan-enriched cubosomes can be regarded as a promising carrier for safe and effective topical ocular delivery.

Designing a green-emitting viscosity-sensitive 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) probe for plasma membrane viscosity imaging.

Viscosity is a key characteristic of lipid membranes - it governs the passive diffusion of solutes and affects the lipid raft formation and membrane fluidity. Precise determination of viscosity values in biological systems is of great interest and viscosity-sensitive fluorescent probes offer a convenient solution for this task. In this work we present a novel membrane-targeting and water-soluble viscosity probe BODIPY-PM, which is based on one of the most frequently used probes BODIPY-C10. Despite its regular use, BODIPY-C10 suffers from poor integration into liquid-ordered lipid phases and lack of water solubility. Here, we investigate the photophysical characteristics of BODIPY-PM and demonstrate that solvent polarity only slightly affects the viscosity-sensing qualities of BODIPY-PM. In addition, with fluorescence lifetime imaging microscopy (FLIM), we imaged microviscosity in complex biological systems - large unilamellar vesicles (LUVs), tethered bilayer membranes (tBLMs) and live lung cancer cells. Our study showcases that BODIPY-PM preferentially stains the plasma membranes of live cells, equally well partitions into both liquid-ordered and liquid-disordered phases and reliably distinguishes lipid phase separation in tBLMs and LUVs.

Biokompatibilni lipidni nanonosači za poboljšanu isporuku fluocinolonacetonida u kožu - fizičkohemijske osobine i in vitro učinak

In order to improve the delivery of topical corticosteroids into inflammatory skin lesions while reducing the likelihood of adverse effects, lipid nanocarriers have received increasing attention. Hence, this study aimed to develop biocompatible nanoemulsions (NEs) and nanostructured lipid carriers (NLCs) as carriers for fluocinolone acetonide (FA) by carefully optimizing the formulation and process parameters. After an analysis of the relevant physicochemical parameters and stability testing, in vitro release and permeation tests were performed to evaluate whether the nanocarriers affected the penetration of FA into/through the skin compared to a conventional reference product (Sinoderm® cream). The developed NEs exhibited satisfactory physicochemical properties (droplet size <200 nm, PDI<0.2, ZP>|-30| mV, pH ~ 4.75) and long-term stability. Although the developed NLCs initially had satisfactory properties, gelation was observed within 3 months of storage, implying that further formulation testing is required to resolve the limited stability of these systems. In vitro release/permeation findings suggest that the developed nanocarriers (especially NEs) provide better delivery of FA into/though the skin compared to the Sinoderm® cream. Therefore, a lecithin-based NE with a 10% lipid phase (medium-chain triglycerides/oleic acid 3:1) is a promising strategy for improved delivery of FA to the inflamed skin, allowing for ease of application, especially to larger skin surfaces and hairy regions.

Approach1es for evolutionary, biochemical, and structural analysis of bacterial steroid 5α-reductases.

Steroid 5α-reductases (SRD5As), also known as 3-oxo-5α-steroid 4-dehydrogenases, are essential membrane-bound enzymes involved in steroid metabolism. Belonging to the NADPH-dependent oxidoreductase family, 5α-reductases catalyze steroids with 3-oxo-Δ4 structure, such as testosterone or progesterone, to produce their corresponding 3-oxo-5α steroids, which are necessary for a variety of physiological and pathological activities. Despite their significance, SRD5A structures are still in short supply to date. Here we describe a protocol for expression, purification, crystallization, structural determination, and functional analysis of PbSRD5A, the 5α-reductase from Proteobacteria bacterium sharing high sequence identity with human SRD5A1 and SRD5A2 isozymes, which we have recently structurally characterized using a lipidic cubic phase approach. Application of similar methods to other 5α-reductase isozymes will lead to breakthroughs in the understanding of the structure, function, and mechanism of oxidoreductases implicated in steroid metabolism.

From angular to round: in depth interfacial analysis of binary phosphatidylethanolamine mixtures in the inverse hexagonal phase.

Packing stress in the lipidic inverse hexagonal HII phase arises from the necessity of the ideally cylinder-shaped micelles to fill out the hexagonally-shaped Wigner-Seitz unit cell. Thus, hydrocarbon chains stretch towards the corners and compress in the direction of the flat side of the hexagonal unit cell. Additionally, the lipid/water interface deviates from being perfectly circular. To study this packing frustration in greater detail, we have doped 1-palmitoyl-2-oleoyl-sn-phosphatidylethanolamine (POPE) with increasing molar concentrations of 1,2-palmitoyl-sn-phosphatidylethanolamine (DPPE: 0 to 15 mol%). Due to its effectively longer hydrophobic tails, DPPE tends to aggregate in the corner regions of the unit cell, and thus, increases the circularity of the lipid/water interface. From small angle X-ray diffraction (SAXD) we determined electron density maps. Using those, we analysed the size, shape and homogeneity of the lipid/water interface as well as that of the methyl trough region. At 6 and 9 mol% DPPE the nanotubular water core most closely resembles a circle; further to this, in comparison to its neighbouring concentrations, the 9 mol% DPPE sample has the smallest water core area and smallest number of lipids per circumference, best alleviating the packing stress. Finally, a three-water layer model was applied, discerning headgroup, perturbed and free water, demonstrating that the hexagonal phase is most stable in the direction of the flat faces (compression zones) and least stable towards the vertices of the unit cell (decompression zones).

Cr(vi) permanently binds to the lipid bilayer in an inverted hexagonal phase throughout the reduction process.

Cr(vi) is a harmful, carcinogenic agent with a high permeability rate throughout the lipid membranes. In an intracellular environment and during interactions with cellular membranes, it undergoes an instant reduction to lower oxidation states throughout radical states, recognized as the most dangerous factor for cells. The cellular membrane is the most visible cellular organelle in the interior and exterior of a cell. In this study, liposomes and non-lamellar inverted hexagonal phase lipid structures based on phosphoethanolamine (PE) were used as model cellular bilayers because of their simple composition, preparation procedure, and the many other properties of natural systems. The lipid membranes were subjected to 0.075 mM Cr(vi) for 15 min, after which the Cr content was removed via dialysis. This way, the remaining Cr content could be studied qualitatively and quantitatively. Using the combined XRF/XAS/EPR approach, we revealed that some Cr content (Cr(iii) and Cr(vi)) was still present in the samples even after long-term dialysis at a temperature significantly above the phase transition for the chosen liposome. The amount of bound Cr increased with increasing PE and -C[double bond, length as m-dash]C- bond content in lipid mixtures. Internal membrane order decreased in less fluid membranes, while in more liquified ones, internal order was only slightly changed after subjecting them to the Cr(vi) agent. The results suggest that the inverted hexagonal phase of lipid structures is much more sensitive to oxidation than the lamellar lipid phase, which can play an important role in the strong cytotoxicity of Cr(vi).

Stability and bioaccessibility of α-tocopherol-enriched nanoemulsions containing different edible oils as carriers

Nanoemulsions have been described as carriers of bioactive compounds since they can protect them during digestion and improve their bioaccessibility. However, their stability and bioaccessibility can be influenced by nanoemulsion composition, mainly by the lipid phase characteristics. Consequently, this study aimed to evaluate three different carrier oils (avocado-AVO, flaxseed-FLA, and chia-CHI) on the stability and bioaccessibility of α-tocopherol carried in nanoemulsions. Particle characteristics (particle size-PS and polydispersity index-PdI) and oxidative stability of nanoemulsions, and α-tocopherol degradation during storage were determined. In addition, free fatty acids (FFAs) release kinetics and α-tocopherol bioaccessibility during in vitro digestion were analyzed. Results showed that all nanoemulsions were colloidally stable due to their droplet characteristics (PS < 200 nm and PdI<0.2), where no phase separation was observed. AVO nanoemulsion was the most stable against oxidation due to its high monounsaturated and saturated fatty acids, while CHI nanoemulsion showed the highest chemical stability of α-tocopherol during storage. Besides, AVO nanoemulsion showed the most significant release of FFA (94%) and the highest α-tocopherol bioaccessibility (81%). In conclusion, the oil type affects the chemical stability and bioaccessibility of α-tocopherol carried in nanoemulsions. Thus, AVO oil can be used as a carrier of fat-soluble vitamins to increase its bioaccessibility during digestion.

Natamycin interferes with ergosterol-dependent lipid phases in model membranes

Natamycin is an antifungal polyene macrolide that is used as a food preservative but also to treat fungal keratitis and other yeast infections. In contrast to other polyene antimycotics, natamycin does not form ion pores in the plasma membrane, but its mode of action is poorly understood. Using nuclear magnetic resonance (NMR) spectroscopy of deuterated sterols, we find that natamycin slows the mobility of ergosterol and cholesterol in liquid-ordered (Lo) membranes to a similar extent. This is supported by molecular dynamics (MD) simulations, which additionally reveal a strong impact of natamycin dimers on sterol dynamics and water permeability. Interference with sterol-dependent lipid packing is also reflected in a natamycin-mediated increase in membrane accessibility for dithionite, particularly in bilayers containing ergosterol. NMR experiments with deuterated sphingomyelin (SM) in sterol-containing membranes reveal that natamycin reduces phase separation and increases lipid exchange in bilayers with ergosterol. In ternary lipid mixtures containing monounsaturated phosphatidylcholine, saturated SM, and either ergosterol or cholesterol, natamycin interferes with phase separation into Lo and liquid-disordered (Ld) domains, as shown by NMR spectroscopy. Employing the intrinsic fluorescence of natamycin in ultraviolet-sensitive microscopy, we can visualize the binding of natamycin to giant unilamellar vesicles (GUVs) and find that it has the highest affinity for the Lo phase in GUVs containing ergosterol. Our results suggest that natamycin specifically interacts with the sterol-induced ordered phase, in which it disrupts lipid packing and increases solvent accessibility. This property is particularly pronounced in ergosterol containing membranes, which could underlie the selective antifungal activity of natamycin.

Simulated dynamic cholesterol redistribution favors membrane fusion pore constriction

Endo- and exocytosis proceed through a highly strained membrane fusion pore topology regardless of the aiding protein machinery. The membrane’s lipid components bias fusion pores toward expansion or closure, modifying the necessary work done by proteins. Cholesterol, a key component of plasma membranes, promotes both inverted lipid phases with concave leaflets (i.e., negative total curvature, which thins the leaflet) and flat bilayer phases with thick, ordered hydrophobic interiors. We demonstrate by theory and simulation that both leaflets of nascent catenoidal fusion pores have negative total curvature. Furthermore, the hydrophobic core of bilayers with strong negative Gaussian curvature is thinned. Therefore, it is an open question whether cholesterol will be enriched in these regions because of the negative total curvature or depleted because of the membrane thinning. Here, we compare all-atom molecular dynamics simulations (built using a procedure to create specific fusion pore geometries) and theory to understand the underlying reasons for lipid redistribution on fusion pores. Our all-atom molecular dynamics simulations resolve this question by showing that cholesterol is strongly excluded from the thinned neck of fusion and fission pores, revealing that thickness (and/or lipid order) influences cholesterol distributions more than curvature. The results imply that cholesterol exclusion can drive fusion pore closure by creating a small, cholesterol-depleted zone in the neck. This model agrees with literature evidence that membrane reshaping is connected to cholesterol-dependent lateral phase separation.

Encapsulation and release of hydrocortisone from proliposomes govern vaginal delivery.

Topical preparations of hydrocortisone can be used for the anti-inflammatory treatment of the female genital area. Although the drug is a low-strength corticosteroid, systemic absorption and distribution of the drug are the most common safety risks associated with this therapy. In the current investigation, we elucidate the physicochemical properties of lipid-based drug carrier systems that govern the local bioavailability of hydrocortisone for intravaginal administration. For this purpose, we compared various proliposome formulations with a commercial cream. Depending on the availability of physiological acceptors, encapsulation and drug release from the lipid phase were found to be the most important drivers of drug bioavailability. The high permeability of hydrocortisone leads to rapid transport of the drug across the mucosal cell layer as indicated by experiments using HEC-1-A and CaSki cell monolayer models. Under sink conditions, differences in the release from the liposomes as determined in the Dispersion Releaser were almost negligible. However, under non-sink conditions, the drug release plateaued at levels corresponding to the encapsulation efficiency. After redispersion, all liposomal formulations performed better than the commercial drug product indicating that the encapsulation into the lipid phase is the main driver sustaining the release.

A Mechanism of Double-Membrane Vesicle Formation from Liquid-Ordered/Liquid-Disordered Phase Separated Spherical Membrane.

Genome replication of coronaviruses takes place in specific cellular compartments, in so-called double-membrane vesicles (DMVs), formed from the endoplasmic reticulum (ER). An intensive production of DMVs is induced by non-structural viral proteins. Here, we proposed a possible mechanism of the DMV formation from ER-derived spherical vesicles where liquid-ordered and liquid-disordered lipid phases coexist. These vesicles are supposed to divide into two homogeneous liquid-ordered and liquid-disordered vesicles. The formation of two spherical vesicles constituting DMV requires a mechanical work to be performed. We considered the excess energy of the boundary between the coexisting lipid phases as the main driving force behind the division of the initial vesicle. Explicitly accounting for the energy of elastic deformations and the interphase boundary energy, we analyzed a range of physical parameters where the DMV formation is possible. We concluded that this process can principally take place in a very narrow range of system parameters. The most probable diameter of DMVs formed according to the proposed mechanism appeared to be approximately 220 nm, in an agreement with the average diameter of DMVs observed in vivo. Our consideration predicts the DMV size to be strongly limited from above. The developed analysis can be utilized for the production of DMVs in model systems.