RESUMO
Glycosaminoglycans (GAGs) are essential components of the extracellular matrices (ECMs) located on the outer surface of cellular membranes. They belong to the group of polysaccharides involved in diverse biological processes acting on the surface and across natural lipid membranes. Recently, particular attention has been focused on possible role of GAGs in the amyloid deposits. The amyloid formation is related to a disorder in protein folding, causing that soluble-in normal conditions-peptides become deposited extracellularly as insoluble fibrils, impairing tissue structure and its function. One of the hypothesis holds that GAGs may inhibit amyloid formation by interacting with the lipid membrane by blocking the accumulation of protein aggregates on the membrane surface. Although the biophysical properties of GAGs are described rather well, little is known about the nature of association between these polysaccharides and components of natural cell membranes. Therefore, a study of GAGs influence on membrane lipids is of particular importance. The aim of the present work is to get insight into the effect of hydrophilic dextran sulfate (DS)-that can be considered as GAG analogue-on membrane lipids organization. This study was based on examining interactions between DS sodium salt of molecular weight equal to about 40 kDa (DS40), dissolved in water subphase, and a model membrane, mimicked as Langmuir monolayer, formed by representative natural membrane lipids: cholesterol and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) as well as their mixtures. Due to the fact that calcium ions in excess may accumulate in the lipid membrane, attracting high molecular weight molecules to their surface, the influence of calcium ions present in the subphase on the DS40 activity has also been examined. It has been found that negatively charged DS, forming a sublayer underneath the monolayer, barely interacts with membrane lipids; however, in the presence of calcium ions the electrostatic interactions between DS40 and lipid membrane are significantly enhanced, leading to the formation of network-like crystalline structures at the surface of model membrane, which can prevent incorporation and interaction with other extracellular molecules, e.g., proteins.
Assuntos
1,2-Dipalmitoilfosfatidilcolina/química , Colesterol/química , Sulfato de Dextrana/química , Glicosaminoglicanos/química , Lipídeos de Membrana/química , Interações Hidrofóbicas e Hidrofílicas , Bicamadas Lipídicas/química , Membranas ArtificiaisRESUMO
Oxysterols are products of cholesterol oxidation. They can be formed endogenously (in both enzymatic and non-enzymatic reactions) as well as exogenously (delivered with food). Recent studies clearly demonstrate cytotoxic properties of these compounds, being mainly due to their incorporation into natural lipid bilayers. This process can influence mechanical and physicochemical properties of biomembrane-mainly by modifying the interactions between its components, which may result in the disruption of proper functioning of cell membrane and could lead to its degradation. Therefore, it can be assumed that oxysterols may affect the initiation of neurodegenerative diseases, including Alzheimer's disease. However, the mode of action of these molecules at the molecular level is not fully known. To get a better understanding of the role of oxysterols in neurodegeneration, it is of great importance to examine mutual interactions between oxysterols and neuronal membrane components. One of the most promising techniques that can be used to analyze such interactions is the Langmuir monolayer technique. In this work, we have prepared an artificial neuronal membrane modeled as multicomponent Langmuir monolayer built up with cholesterol, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and sphingomyelin (SM). To examine whether there are any changes in the membrane properties under oxidative stress, in this paper we have investigated the impact of the representative ring-oxidized oxysterol: 7-ketocholesterol (7-KC). Our results show that replacing cholesterol with 7-KC increases the interaction between molecules in the model membrane.
Assuntos
Membrana Celular/química , Cetocolesteróis/química , Bicamadas Lipídicas/química , Modelos Químicos , Neurônios/química , Membrana Celular/metabolismo , Cetocolesteróis/metabolismo , Bicamadas Lipídicas/metabolismo , Neurônios/metabolismo , Fosfatidilcolinas/química , Fosfatidilcolinas/metabolismo , Esfingomielinas/química , Esfingomielinas/metabolismoRESUMO
Outer layer of cellular membrane contains ordered domains enriched in cholesterol and sphingolipids, called 'lipid rafts', which play various biological roles, i.e., are involved in the induction of cell death by apoptosis. Recent studies have shown that these domains may constitute binding sites for selected drugs. For example alkylphosphocholines (APCs), which are new-generation antitumor agents characterized by high selectivity and broad spectrum of activity, are known to have their molecular targets located at cellular membrane and their selective accumulation in tumor cells has been hypothesized to be linked with the alternation of biophysical properties of lipid rafts. To get a deeper insight into this issue, interactions between representative APC: erucylphosphocholine, and artificial lipid raft system, modeled as Langmuir monolayer (composed of cholesterol and sphingomyelin mixed in 1:2 proportion) were investigated. The Langmuir monolayer experiments, based on recording surface pressure-area isotherms, were complemented with Brewster angle microscopy results, which enabled direct visualization of the monolayers structure. In addition, the investigated monolayers were transferred onto solid supports and studied with AFM. The interactions between model raft system and erucylphosphocholine were analyzed qualitatively (with mean molecular area values) as well as quantitatively (with ΔG(exc) function). The obtained results indicate that erucylphosphocholine introduced to raft-mimicking model membrane causes fluidizing effect and weakens the interactions between cholesterol and sphingomyelin, which results in phase separation at high surface pressures. This leads to the redistribution of cholesterol molecules in model raft, which confirms the results observed in biological studies.
Assuntos
Antineoplásicos/química , Microdomínios da Membrana/química , Membranas Artificiais , Modelos Químicos , Fosforilcolina/análogos & derivados , Animais , Humanos , Fosforilcolina/químicaRESUMO
Cyclosporin A (CsA), a hydrophobic cyclic peptide produced by the fungus Tolypocladium inflatum, is well known for its high efficiency as an immunosuppressor for transplanted organs and anti-inflammatory properties; however, it is also active as antiparasitic (antimalarial) drug. Antimalarial mechanism of CsA action lacks a detailed understanding at molecular level. Due to a high lipophilicity of CsA, it is able to interact with lipids of cellular membrane; however, molecular targets of this drug are still unknown. To get a deeper insight into the mode of antimalarial activity of CsA, it is of utmost importance to examine its interactions with membrane components. To reach this goal, the Langmuir monolayer technique, which serves as a very useful, easy to handle and controllable model of biomembranes, has been employed. In this work, the interactions between CsA and main membrane lipids, i.e., cholesterol (Chol), 2-oleoyl-1-palmitoyl-3-phosphocholine (POPC), and sphingomyelin (SM), have been investigated. Attractive interactions are observed only for CsA mixtures with SM, while repulsive forces occur in systems containing remaining membrane lipids. Taking into consideration mutual interactions between membrane lipids (Chol-SM; Chol-POPC and SM-POPC), the behavior of CsA in model erythrocyte membrane of normal and infected cells has been analyzed. Our results prove strong affinity of CsA to SM in membrane environment. Since normal and parasitized erythrocytes differ significantly in the level of SM, this phospholipid may be considered as a molecular target for antimalarial activity of CsA.
Assuntos
Antimaláricos/química , Ciclosporina/química , Lipídeos de Membrana/química , Colesterol/química , Membrana Eritrocítica/química , Membrana Eritrocítica/metabolismo , Humanos , Modelos Moleculares , Estrutura Molecular , Fosfatidilcolinas/química , Esfingomielinas/químicaRESUMO
Cyclosporin A (CsA), a hydrophobic peptide, mainly known for its immunosuppressant properties, has shown a broad range of biological activities, including antimalarial action. Since CsA was found to be active on membrane level, it was subjected for investigations involving membrane models. Our former studies on interactions between CsA and different membrane lipids using Langmuir monolayer technique indicated its affinity for sphingomyelin (SM). Inspired by this finding we have extended our experiments on multicomponent systems and performed systematic investigations of CsA behavior towards artificial membranes containing different mutual proportion of sphingomyelin and cholesterol (Chol). Langmuir monolayer results have been complemented with in-situ films structure visualization applying Brewster angle microscopy (BAM) and, after films transfer onto solid support, atomic force microscopy (AFM). Our results show that cyclosporin A introduced to SM:Chol mixed monolayers distributes differently, depending on SM-to-Chol proportion. In raft-mimicking (2:1) stoichiometry, even distribution of the drug within SM:Chol matrix was observed. However, in SM:Chol model membranes of different proportion (3:1; 1:1; 1:2), containing either the excess of unbound sphingomyelin or cholesterol in addition to model lipid raft domains, introduction of CsA induced a phase separation.
Assuntos
Colesterol/química , Ciclosporina/química , Membranas Artificiais , Esfingomielinas/química , Lipídeos de Membrana/químicaRESUMO
A series of triblock semifluorinated n-alkanes of the general formula: F(CF2)m(CH2)n(CF2)mF, (in short FmHnFm), where m=10, 12, and n=6, 8, and 12 have been synthesized and employed for liquid crystalline studies and Langmuir monolayer characterization. Differential scanning calorimetry (DSC) measurements together with texture observation with polarizing microscope (POM) revealed the presence of liquid crystalline smectic phases for all the investigated homologs. The behavior of the studied molecules spread at the free water surface has also been investigated. Our results show for the first time that these unusual film-forming materials, which are completely hydrophobic in nature and do not possess any polar group in their structure, are surface active and form insoluble (Langmuir) monolayers at the air/water interface. Due to the fact that these molecules are chemically inert and, similar to the semifluorinated diblocks, are not toxic, they may be destined for biomedical uses as gas carriers and contrast agents, as well as in drug delivery systems.