Model Lipid Monolayers Research Articles

Cerebrosides lipids and their physical effect on multiple sclerosis membranes.

The myelin sheath is a critical component of the central nervous system (CNS) and peripheral nervous system (PNS). It is a multilamellar membrane consisting of a coiled bilayer that acts as an electrical insulator for nerve impulses traveling along axons between nerve cells. Compared to other membranes in the body, the myelin sheath contains a large lipid fraction in the range of 75-80% of the dry mass, compared to about 50% in other membranes, consisting of many different lipids. The lipids in the myelin sheath play a critical role in membrane stability and compaction. While previous investigations have used model lipid bilayers and monolayers to study their interactions with important proteins including myelin basic protein (MBP) and proteolipid protein (PLP), our understanding of the role of lipid composition on membrane formation thermodynamics and rheology, particularly the effects produced by lipid cerebrosides (Hydroxylated cerebrosides, Non Hydroxylated cerebrosides and Cerebroside sulfatide). In this presentation we study the formation and interfacial dilatational rheology of lipid monolayers. We use a Langmuir trough for both surface pressure versus area isotherms and oscillating area interfacial rheology measurements, all performed at approximately body temperature (37°C). From isotherm data, we calculate differences in the Gibbs free energy of mixing between the four monolayers to show differences in intermolecular interactions and miscibility. These results are used to support observations in interfacial rheology results, where. We also analyze the nonlinear rheological behavior of the monolayers, for example observing in general film softening during compression and expansion. These first oscillatory interfacial rheology measurements of model myelin monolayers, coupled with the thermodynamic analysis of film formation, improve our understanding of the lipid the film properties, and provide clues to how the changes in lipid composition may lead to demyelination.

How do antimicrobial peptides disrupt the lipopolysaccharide membrane leaflet of Gram-negative bacteria?

HypothesisIt is widely regarded that antimicrobial peptides (AMPs) kill bacteria by physically disrupting microbial membranes and causing cytoplasmic leakage, but it remains unclear how AMPs disrupt the outer membrane (OM) of Gram-negative bacteria (GNB) and then compromise the inner membrane. We hypothesise that different AMPs impose different structural disruptions, with direct implications to their antimicrobial efficacies. ExperimentsThe antimicrobial activities of three typical AMPs, including the designed short AMP, G3, and two natural AMPs, melittin and LL37, against E. coli and their haemolytic activities were studied. Lipopolysaccharide (LPS) and anionic di-palmitoyl phosphatidyl glycerol (DPPG) monolayer models were constructed to mimic the outer membrane and inner membrane leaflets of Gram-negative bacteria. The binding and penetration of AMPs to the model lipid monolayers were systematically studied by neutron reflection via multiple H/D contrast variations. FindingG3 has relatively high antimicrobial activity, low cytotoxicity, and high proteolytic stability, whilst melittin has significant haemolysis and LL37 has weaker antimicrobial activity. G3 could rapidly lyse LPS and DPPG monolayers within 10–20 min. In contrast, melittin was highly active against the LPS membrane, but the dynamic process lasted up to 80 min, with excessive stacking in the OM. LL37 caused rather weak destruction to LPS and DPPG monolayers, leading to massive adsorption on the membrane surface without penetrating the lipid tail region. These findings demonstrate that the rationally designed AMP G3 was well optimised to impose most effective destruction to bacterial membranes, consistent with its highest bactericidal activity. These different interfacial structural features associated with AMP binding shed light on the future development of active and biocompatible AMPs for infection and wound treatments.

Effect of the oat, horse chestnut, cowherb, soy, quinoa and soapwort extracts on skin‐mimicking monolayers and cell lines

AbstractThe main goal of the study was to compare the effect of aqueous extracts of oat (Avena sativa L.), horse chestnut (Aesculus hippocastanum L.), quinoa (Chenopodium quinoa Willd.), soapwort (Saponaria officinalis L.), cowherb (Vaccaria hispanica [P. Mill.] Rauschert) and soy (Glycine max L.) on model lipid monolayers mimicking the lipid membrane of keratinocytes and intercellular lipids of stratum corneum, and on human skin‐related cell lines. Two lipid monolayers, consisting of a dipalmitoylphosphatidylcholine (DPPC) and cholesterol mixture in a molar ratio of 7:3 and Ceramide VI, stearic acid and cholesterol in a molar ratio of 1:1:0.7, and two cell lines (human keratinocyte HaCaT and human skin malignant melanoma A375) were employed. None of the extracts reduced surface pressure below the level achieved for bare monolayers. The strength of the effect on the lipid monolayers (horse chestnut > cowherb > soapwort > soy) points to the existence of some specific interactions responsible for the observed affinity of biosurfactants from the extracts to the lipids in the monolayers. The cytotoxicity tests performed with two model skin cell lines showed that all six plants extracts significantly reduced the cells' viability in a concentration‐dependent way. The model lipid monolayers were not solubilized by the investigated surface‐active extracts. The latter thus proved interesting candidates for application in mild cleansing cosmetic formulations. Penetration of the monolayers by surface‐active components of some extracts, especially horse chestnut, cowherb and soapwort, opens new possibilities for topical delivery of active components.

Interaction of Amyloid-β-(1-42) Peptide and Its Aggregates with Lipid/Water Interfaces Probed by Vibrational Sum-Frequency Generation Spectroscopy.

In this study, we use surface-sensitive vibrational sum-frequency generation (VSFG) spectroscopy to investigate the interaction between model lipid monolayers and Aβ(1-42) in its monomeric and aggregated states. Combining VSFG with atomic force microscopy (AFM) and thioflavin T (ThT) fluorescence measurements, we found that only small aggregates with probably a β-hairpin-like structure adsorbed to the zwitterionic lipid monolayer (DOPC). In contrast, larger aggregates with an extended β-sheet structure adsorbed to a negatively charged lipid monolayer (DOPG). The adsorption of small, initially formed aggregates strongly destabilized both monolayers, but only the DOPC monolayer was completely disrupted. We showed that the intensity of the amide-II' band in achiral (SSP) and chiral (SPP) polarization combinations increased in time when Aβ(1-42) aggregates accumulated at the DOPG monolayer. Nevertheless, almost no adsorption of preformed mature fibrils to DOPG monolayers was detected. By performing spectral VSFG calculations, we revealed a clear correlation between the amide-II' signal and the degree of amyloid aggregates (e.g., oligomers or (proto)fibrils) of various Aβ(1-42) structures. The calculations showed that only structures with a significant amyloid β-sheet content have a strong amide-II' intensity, in line with previous Raman studies. The combination of the presented results substantiates the amide-II(') band as a legitimate amyloid marker.

Surface-active extracts from plants rich in saponins – effect on lipid mono- and bilayers

The aqueous extracts of the seeds of oat (Avena sativa L.), horse chestnut (Aesculus hippocastanum L.), soybean (Glycine max L.), cowherb (Vaccaria hispanica [P. Mill.] Rauschert) and quinoa (Chenopodium quinoa Willd.), and the roots of soapwort (Saponaria officinalis L.) without any preservatives were characterized in terms of their surface tension, surface compression (dilational) rheology, foamability and foam stability. The saponin content in the extracts was determined using UPLC-MS and their interaction with model lipid monolayers consisting of dipalmitoylphosphatidylcholine (DPPC)/cholesterol and Ceramide AP/stearic acid/cholesterol were analyzed by surface pressure relaxation, surface compression elasticity and neutron reflectometry (NR). The lipid composition was chosen to mimic the cell membrane of keratinocytes – major constituents of the human deeper skin layers, and the intercellular lipids (“mortar”) in the “bricks and mortar” model of the outermost layer of the epidermis (stratum corneum). Bilayers of DPPC/cholesterol were additionally characterized using dynamic light scattering (DLS) and NR. The oat and soybean extracts were shown to be much less abundant in saponins as compared to cowherb, horse chestnut, soapwort or quinoa, and showed limited foaming abilities. They did not affect significantly the model lipid mono- and bilayers mimicking the skin outer layers, either. The horse chestnut extract affected both model membranes to the highest extent, yet without solubilizing the lipids.

Effect of synthetic surfactants and soapwort (Saponaria officinalis L.) extract on skin-mimetic model lipid monolayers

The effect of a saponin-rich extract from rhizomes of Soapwort (Saponaria officinalis L) and four synthetic surfactants: sodium lauryl sulphate (SLS), sodium laureth sulphate (SLES), ammonium lauryl sulphate (ALS) and cocamidopropyl betaine (CAPB) on two model lipid monolayers is analyzed using surface pressure, surface dilatational rheology and fluorescence microscopy. The following monolayers were employed: dipalmitoylphosphatidylcholine/cholesterol mixture in a molar ratio of 7:3 (DPPC/CHOL) and Ceramide [AP]/stearic acid/cholesterol in a molar ratio of 14:14:10 (CER/SA/CHOL). They mimicked a general bilayer structure and an intercellular lipid mixture, respectively. Both lipid mixtures on Milli-Q water were first compressed to the initial surface pressure, Π0 = 30 mN/m and then the subphase was exchanged with the respective (bio)surfactant solution at 1% (w/w). All four synthetic surfactants behaved in a similar way: they increased surface pressure to about 40 mN/m and reduced the storage modulus of surface dilational surface rheology, E′, to the values close to zero. The corresponding fluorescence microscopy pictures confirmed that the lipids mimicking the stratum corneum components were almost completely removed by the synthetic surfactants under the present experimental conditions. The components of the Soapwort extract (SAP) increased surface pressure to significantly higher values than the synthetic surfactants, but even more spectacular increase was observed for the storage modulus of the SAP-penetrated lipid monolayers (up to E′= 715 mN/m).

Effect of saponins from quinoa on a skin-mimetic lipid monolayer containing cholesterol

The study discusses the effect of a quinoa seed coat extract on a cholesterol-based Langmuir monolayer mimicking the intercellular lipid mixture in the skin’s outermost layer - stratum corneum. Besides cholesterol (CHOL), the monolayer contains also stearic acid (SA) and ceramide VI (CER), in a molar ratio of 10:14:14. Three quinoa extracts were tested for their surface activity: a) from the whole seed, b) from the dehulled seed, and c) from the seed coat. The latter shows significantly higher ability to reduce surface tension (increase surface pressure) than the others. Its adsorbed layers display also reasonable surface dilational elasticity (storage) modulus, E′. These observations are in line with the literature reports on the high concentrations of triterpenoid glycosidic biosurfactants – saponins, in quinoa seed, especially in its coat. The saponin-rich extract of quinoa seed coat was thus introduced underneath the pre-formed lipid monolayer compressed to surface pressure, Π = 30 mN/m in a Langmuir trough, in order to register the surface pressure response. The increase of both the surface pressure and surface dilational elasticity modulus suggests that saponins, and possibly other surface-active components of the extract, incorporate into the model lipid monolayer, without solubilizing it. This opens new perspectives for the saponin-rich quinoa seed extract as skin penetration-enhancing active components for cosmetics or pharmaceutical purposes.

Interaction of Myelin Basic Protein with Myelin-like Lipid Monolayers at Air-Water Interface.

Interaction of myelin basic protein (MBP) and the cytoplasmic leaflets of the oligodendrocyte membrane is essential for the formation and compaction of the myelin sheath of the central nervous system and is altered aberrantly and implicated in the pathogenesis of neurodegenerative diseases like multiple sclerosis. To gain more detailed insights into this interaction, the adsorption of MBP to model lipid monolayers of similar composition to the myelin of the central nervous system was studied at the air-water interface with monolayer adsorption experiments. Measuring the surface pressure and the related maximum insertion pressure of MBP for different myelin-like lipid monolayers provided information about the specific role of each of the single lipids in the myelin. Depending on the ratio of negatively charged lipids to uncharged lipids and the distance between charges, the adsorption process was found to be determined by two counteracting effects: (i) protein incorporation, resulting in an increasing surface pressure and (ii) lipid condensation due to electrostatic interaction between the positively charged protein and negatively charged lipids, resulting in a decreasing surface pressure. Although electrostatic interactions led to high insertion pressures, the associated lipid condensation lowered the fluidity of the myelin-like monolayer.

Interaction of thermal responsive NIPAM nanogels with model lipid monolayers at the air-water interface

Understanding the interaction of nanoparticles (NP) with ceramide lipids is important in developing strategies to overcome the formidable obstacle that is skin. This paper presents studies of interactions between N-isopropylacrylamide nanogels, crosslinked with 30% N,N′-methylenebisacrylamide, and model ceramide lipid monolayers at the air-water interface as a function of temperature. In the case of the mixed ceramide/cholesterol/behenic acid monolayer, the interaction of nanogels with the ceramide was strongly mediated by the fatty acids. This interaction between nanogels and monolayer components is dominated by hydrophobic-hydrophobic binding. The data show the important intermediary role of the fatty acid in facilitating transmembrane transport. For a pure ceramide lipid monolayer, the neutron reflectivity (NR), Brewster angle microscopy (BAM) and surface pressure results showed a lipid-nanogel complex formation and the subsequent depletion/solubilisation of the lipids from the interface when the area per molecule for the lipid was increased from 42 to 44 Å2.

Antimicrobial activity of lysozyme isoforms: Key molecular features.

Increasing bacterial resistance towards antibiotics has stimulated research for novel antimicrobials. Proteins acting on bacterial membranes could be a solution. Lysozyme has been proven active against E. coli by disruption of both outer and cytoplasmic membranes, with dry-heating increasing lysozyme activity. Dry-heated lysozyme (DH-L) is a mixture of isoforms (isoaspartyl, native-like and succinimide lysozymes), giving rise to two questions: what effects does each form have, and which physicochemical properties are critical as regards the antibacterial activity? These issues were investigated by fractionating DH-L, analyzing structural properties of each fraction, and testing each fraction in vivo on bacteria and in vitro on membrane models. Positive net charge, hydrophobicity and molecular flexibility of the isoforms seem key parameters for their interaction with E. coli membranes. The succinimide lysozyme fraction, the most positive, flexible and hydrophobic, shows the highest antimicrobial activity, induces the strongest bacterial membrane disruption and is the most surface active on model lipid monolayers. Moreover, each fraction appears less efficient than DH-L against E. coli, indicating a synergetic cooperation between lysozyme isoforms. The bacterial membrane modifications induced by one isoform could facilitate the subsequent action of the other isoforms.

Ballistic Penetration of Highly Charged Nanoaerosol Particles through a Lipid Monolayer.

To be used as a drug, inhaled nanoaerosol particles (NAPs) must first penetrate the lipid layer on top of the lung fluid before they will be able to reach the lung epithelium. We investigated how the penetration of NAPs through a model lipid monolayer (LM) depends upon their charging level and size. It was shown that deposition of NAPs 20-200 nm in diameter and charged to the Rayleigh limit gradually increased the surface tension of a dipalmitoylphosphatidylcholine monolayer (DPPC), indicating a loss of lipid molecules from the monolayer. This phenomenon was reproduced with a variety of NAPs produced from glucose, proteins, and polymers. Transfer of the lipid material into the subphase was documented by direct visualization of lipid nanoparticles in the subphase with atomic force microscopy after deposition of glucose NAPs on a DPPC monolayer, followed by collection of the lipid nanoparticles on a mica surface. Partial restoration of tension upon storage indicates that some of the lipid may return to the monolayer. Experiments with the deposition of highly charged calibrated polystyrene nanoparticles showed that the amount of lipid removed from the surface was roughly proportional to the overall surface area of the deposited NAPs. When the number of charges on the NAPs was reduced from their Rayleigh level of 103-104 units to 1-10 units, no notable changes in monolayer surface tension were observed even with prolonged deposition of such NAPs. It was therefore concluded that only highly charged NAPs of a certain size acquire sufficient speed from their attraction by mirror charges to enable ballistic penetration through a lipid monolayer.

Interaction of acylated and unacylated forms of E. coli alpha-hemolysin with lipid monolayers: a PM-IRRAS study

Uropathogenic strains of Escherichia coli produce virulence factors, such as the protein toxin alpha-hemolysin (HlyA), that enable the bacteria to colonize the host and establish an infection. HlyA is synthetized as a protoxin (ProHlyA) that is transformed into the active form in the bacterial cytosol by the covalent linkage of two fatty-acyl moieties to the polypeptide chain before the secretion of HlyA into the extracellular medium. The aim of this work was to investigate the effect of the fatty acylation of HlyA on protein conformation and protein-membrane interactions. Polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS) experiments were performed at the air-water interface, and lipid monolayers mimicking the outer leaflet of red-blood-cell membranes were used as model systems for the study of protein-membrane interaction. According to surface-pressure measurements, incorporation of the acylated protein into the lipid films was faster than that of the nonacylated form. PM-IRRAS measurements revealed that the adsorption of the proteins to the lipid monolayers induced disorder in the lipid acyl chains and also changed the elastic properties of the films independently of protein acylation. No significant difference was observed between HlyA and ProHlyA in the interaction with the model lipid monolayers; but when these proteins became adsorbed on a bare air-water interface, they adopted different secondary structures. The assumption of the correct protein conformation at a hydrophobic-hydrophilic interface could constitute a critical condition for biologic activity.

Identifying the selectivity of antimicrobial peptides to cell membranes by sum frequency generation spectroscopy.

Cationic amphiphilic peptides have been engineered to target both Gram-positive and Gram-negative bacteria while avoiding damage to other cell types. However, the exact mechanism of how these peptides target, bind, and disrupt bacterial cell membranes is not understood. One specific peptide that has been engineered to selectively capture bacteria is WLBU2 (sequence: RRWVRRVRRWVRRVVRVVRRWVRR). It has been suggested that WLBU2 activity stems from the fact that when interacting with bacterial cell membranes the peptide assumes an α-helical structure and inserts itself into the membrane. Alternatively, in the presence of mammalian cell membranes, the peptide assumes an inert β-sheet structure. To test this hypothesis, the authors applied sum frequency generation (SFG) spectroscopy and surface tensiometry to identify the structure of WLBU2 as it interacts with model lipid monolayers that mimic mammalian and bacterial cell membranes. Model mammalian cell membranes were built upon zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine lipids while bacterial cell membranes were constructed with negatively charged 1,2-dimyristoyl-sn-glycero-3-phospho-(1'-rac-glycerol) lipids. Observed changes in surface pressure at the peptide-lipid-air interface demonstrate that the peptide has a clear binding preference toward negatively charged bacteria-like lipids. The structure of both the lipids and peptides were characterized by SFG spectra collected at the monolayer interface. Changes in monolayer structure as the peptide binds were observed by tracking the intensities of SFG vibrational modes related to the acyl chains within the lipids. Peptide structures when bound to both types of lipids were determined by SFG spectra collected within the amide I vibrational band. The SFG spectra of WLBU2 interacting with the model mammalian lipid monolayer contain two peaks near 1642 and 1678 cm-1 indicative of an inactive β-sheet structure. SFG spectra collected from the peptide bound to a bacteria-like lipid monolayer contains just a single peak near 1651 cm-1 which corresponds to an active α-helix structure. Combined, the tensiometry and SFG results demonstrate that WLBU2 both possesses a higher binding affinity toward and is in an active α-helix structure when bound to bacterial cell membranes.

How charge distribution influences the function of membrane-active peptides: Lytic or cell-penetrating?

Lytic and cell-penetrating peptides (CPPs) are both membrane-active peptides sharing similar physicochemical properties. Although their respective functions have been intensively investigated, the difference of intrinsic properties between these two types of peptides is rarely discussed. In this study, we designed a series of analogs of a recently discovered CPP ZXR-1 (FKIGGFIKKLWRSKLA) by varying the charge distributions both on the helical wheel projection and along the sequence. These peptides showed different functions on cell membranes, including membrane lytic (peptide Z1), cell-penetrating (peptide ZXR-1, Z2 and Z3), and inactive (peptide Z4) peptides. The three groups of peptides displayed different interactions with model lipid monolayer, and found that peptide insertion might be an important dynamic step to distinguish lytic and cell penetrating functions. Based on the analysis of charge distribution patterns, it was proposed that the charge distributions on the helical wheel and along the sequence are both able to influence the functions of the membrane-active peptides. This finding provides a further understanding about the effect of charge distribution on the functions of membrane-active peptides, and will be helpful for the design of functional peptides.

Analysis of the induction of the myelin basic protein binding to the plasma membrane phospholipid monolayer**Project supported by the National Natural Science Foundation of China (Grant Nos. 21402114 and 11544009), the Natural Science Basic Research Plan in Shaanxi Province of China (Grant No. 2016JM2010), the Fundamental Research Funds for the Central Universities of China (Grant No. GK201604004), and the National

Myelin basic protein (MBP) is an essential structure involved in the generation of central nervous system (CNS) myelin. Myelin shape has been described as liquid crystal structure of biological membrane. The interactions of MBP with monolayers of different lipid compositions are responsible for the multi-lamellar structure and stability of myelin. In this paper, we have designed MBP-incorporated model lipid monolayers and studied the phase behavior of MBP adsorbed on the plasma membrane at the air/water interface by thermodynamic method and atomic force microscopy (AFM). By analyzing the pressure–area (π–A) and pressure–time (π–T) isotherms, univariate linear regression equation was obtained. In addition, the elastic modulus, surface pressure increase, maximal insertion pressure, and synergy factor of monolayers were detected. These parameters can be used to modulate the monolayers binding of protein, and the results show that MBP has the strongest affinity for 1,2-dipalmitoyl-sn-glycero-3- phosphoserine (DPPS) monolayer, followed by DPPC/DPPS mixed and 1,2-dipalmitoyl-sn-glycero-3-phospho-choline (DPPC) monolayers via electrostatic and hydrophobic interactions. AFM images of DPPS and DPPC/DPPS mixed monolayers in the presence of MBP (5 nM) show a phase separation texture at the surface pressure of 20 mN/m and the incorporation of MBP put into the DPPC monolayers has exerted a significant effect on the domain structure. MBP is not an integral membrane protein but, due to its positive charge, interacts with the lipid head groups and stabilizes the membranes. The interaction between MBP and phospholipid membrane to determine the nervous system of the disease has a good biophysical significance and medical value.

Atomic force microscopy as a tool to study the adsorption of DNA onto lipid interfaces.

The Atomic Force Microscopy (AFM) technique appears as a central tool for the characterization of DNA adsorption onto lipid interfaces. Regardless of the huge number of surveys devoted to this issue, there are still fascinating phenomena in this field that have not been explored in detail by AFM. For instance, adsorption of DNA onto like-charged lipid surfaces mediated by cations is still not fully understood even though it is gaining popularity nowadays in gene therapy and nanotechnology. Studies related to the complexation of DNA with anionic lipids as a non-viral gene delivery vehicle as well as the formation of self-assembled nanoscale DNA constructs (DNA origami) are two of the most attractive systems. Unfortunately, molecular mechanisms underlying the adsorption of DNA onto anionic lipid interfaces remain unclear so far. In view of that, AFM becomes an appropriate technique to provide valuable information to understand the adsorption of DNA to anionic lipid surfaces. As a second part of this review we provide an illustrative example of application of the AFM technique to probe the DNA adsorption onto a model lipid monolayer negatively charged. Microsc. Res. Tech. 80:11-17, 2017. © 2016 Wiley Periodicals, Inc.

Adsorption of gastric lipase onto multicomponent model lipid monolayers with phase separation

The enzymatic lipolysis of complex natural lipoproteic assemblies such as milk fat globules is central in neonatal digestion. This process first requires the rapid adsorption of a lipolytic enzyme, gastric lipase, onto the membrane enveloping the triglyceride substrate before the onset of catalytic activity. The interactions governing lipase adsorption onto this complex lipid/water interface are not fully elucidated. This study was designed to unravel the interactions of recombinant dog gastric lipase (rDGL) with model monolayers presenting liquid–liquid phase coexistence and mimicking the outer leaflet of the milk fat globule membrane. Combining biophysical tools (ellipsometry, tensiometry and atomic force microscopy), it was evidenced that rDGL partitions toward liquid expanded phase and at phase boundaries. rDGL gets adsorbed at several levels of insertion suggesting molecular cooperation that may favor insertion and strongly impacts on the lipid phase lateral organization. The addition of phosphatidylserine, negatively charged, reinforced adsorption; hence besides hydrophobic interactions and as further investigated through surface potential modeling, rDGL adsorption is favored by electrostatic interactions.

A new tribological experimental setup to study confined and sheared monolayers.

We have developed an original experimental setup, coupling tribology, and velocimetry experiments together with a direct visualization of the contact. The significant interest of the setup is to measure simultaneously the apparent friction coefficient and the velocity of confined layers down to molecular scale. The major challenge of this experimental coupling is to catch information on a nanometer-thick sheared zone confined between a rigid spherical indenter of millimetric radius sliding on a flat surface at constant speed. In order to demonstrate the accuracy of this setup to investigate nanometer-scale sliding layers, we studied a model lipid monolayer deposited on glass slides. It shows that our experimental setup will, therefore, help to highlight the hydrodynamic of such sheared confined layers in lubrication, biolubrication, or friction on solid polymer.

Complexation of phospholipids and cholesterol by triterpenic saponins in bulk and in monolayers

The interactions between three triterpene saponins: α-hederin, hederacoside C and ammonium glycyrrhizate with model lipids: cholesterol and dipalmitoylphosphatidylcholine (DPPC) are described. The oleanolic acid-type saponins (α-hederin and hederacoside C) were shown to form 1:1 complexes with lipids in bulk, characterized by stability constants in the range (4.0±0.2)·103–(5.0±0.4)·104M−1. The complexes with cholesterol are generally stronger than those with DPPC. On the contrary, ammonium glycyrrhizate does not form complexes with any of the lipids in solution. The saponin–lipid interactions were also studied in a confined environment of Langmuir monolayers of DPPC and DPPC/cholesterol with the saponins present in the subphase. A combined monolayer relaxation, surface dilational rheology, fluorescence microscopy and neutron reflectivity (NR) study showed that all three saponins are able to penetrate pure DPPC and mixed DPPC/cholesterol monolayers. Overall, the effect of the saponins on the model lipid monolayers does not fully correlate with the lipid–saponin complex formation in the homogeneous solution. The best correlation was found for α-hederin, for which even the preference for cholesterol over DPPC observed in bulk is well reflected in the monolayer studies and the literature data on its membranolytic activity. Similarly, the lack of interaction of ammonium glycyrrhizate with both lipids is evident equally in bulk and monolayer experiments, as well as in its weak membranolytic activity. The combined bulk and monolayer results are discussed in view of the role of confinement in modulating the saponin–lipid interactions and possible mechanism of membranolytic activity of saponins.

Cholesterol and Sphingomyelin-Containing Model Condensed Lipid Monolayers: Heterogeneities Involving Ordered Microdomains Assessed by Two Cholesterol Derivatives.

Lipid monolayers are often considered as model membranes, but they are also the physiologic lipid part of the peripheral envelope of lipoproteins and cytosolic lipid bodies. However, their structural organization is still rather elusive, in particular when both cholesterol and sphingomyelin are present. To investigate such structural organization of hemimembranes, we measured, using alternative current voltammetry, the differential capacitance of condensed phosphatidylcholine-based monolayers as a function of applied potential, which is sensitive to their lipid composition and molecular arrangement. Especially, monolayers containing both sphingomyelin and cholesterol, at 15% w/w, presented specific characteristics of the differential capacitance versus potential curves recorded, which was indicative of specific interactions between these two lipid components. We then compared the behavior of two cholesterol derivatives (at 15% w/w), 21-methylpyrenyl-cholesterol (Pyr-met-Chol) and 22-nitrobenzoxadiazole-cholesterol (NBD-Chol), with that of cholesterol when present in model monolayers. Indeed, these two probes were chosen because of previous findings reporting opposite behaviors within bilayer membranes regarding their interaction with ordered lipids, with only Pyr-met-Chol mimicking cholesterol well. Remarkably, in monolayers containing sphingomyelin or not, Pyr-met-Chol and NBD-Chol presented contrasting behaviors, and Pyr-met-Chol mimicked cholesterol only in the presence of sphingomyelin. These two observations (i.e., optimal amounts of sphingomyelin and cholesterol, and the ability to discriminate between Pyr-met-Chol and NBD-Chol) can be interpreted by the existence of heterogeneities including ordered patches in sphingomyelin- and cholesterol-containing monolayers. Since such monolayer lipid arrangement shares some properties with the raft-type lipid microdomains well-described in sphingomyelin- and cholesterol-containing bilayer membranes, our data thus strongly suggest the existence of compact and ordered microdomains in model lipid monolayers.