RESUMO
Vitamin D3 deficiency has serious health consequences, as demonstrated by its effect on severity and recovery after COVID-19 infection. Because of high hydrophobicity, its absorption and subsequent redistribution throughout the body are inherently dependent on the accompanying lipids and/or proteins. The effective oral vitamin D3 formulation should ensure penetration of the mucus layer followed by internalization by competent cells. Isothermal titration calorimetry and computer simulations show that vitamin D3 molecules cannot leave the hydrophobic environment, indicating that their absorption is predominantly driven by the digestion of the delivery vehicle. In the clinical experiment, liposomal vitamin D3 was compared to the oily formulation. The results obtained show that liposomal vitamin D3 causes a rapid increase in the plasma concentration of calcidiol. No such effect was observed when the oily formulation was used. The effect was especially pronounced for people with severe vitamin D3 deficiency.
Assuntos
COVID-19 , Colecalciferol , Disponibilidade Biológica , Humanos , LipossomosRESUMO
The molecular details of the passive water flux across the hydrophobic membrane interior are still a matter of debate. One of the postulated mechanisms is the spontaneous, water-filled pore opening, which facilitates the hydrophilic connection between aqueous phases separated by the membrane. In the paper, we provide experimental evidence showing that the spontaneous lipid pore formation correlates with the membrane mechanics; hence, it depends on the composition of the lipid bilayer and the concentration of the osmotically active compound. Using liposomes as an experimental membrane model, osmotically induced water efflux was measured with the stopped-flow technique. Shapes of kinetic curves obtained at low osmotic pressure differences are interpreted in terms of two events: the lipid pore opening and water flow across the aqueous channel. The biological significance of the dependence of the lipid pore formation on the concentration difference of an osmotically active compound was illustrated by the demonstration that osmotically driven water flow can be accompanied by the dissipation of the pH gradient. The application of the Helfrich model to describe the probability of lipid pore opening was validated by demonstrating that the probability of pore opening correlates with the membrane bending rigidity. The correlation was determined by experimentally derived bending rigidity coefficients and probabilities of lipid pores opening.
Assuntos
Membrana Celular/química , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Água/metabolismo , Ovos , Concentração de Íons de Hidrogênio , Cinética , Luz , Lipossomos/química , Lisofosfatidilcolinas/química , Lipídeos de Membrana/química , Modelos Químicos , Pressão Osmótica , Fosfatidilcolinas/química , Fosfatidiletanolaminas/química , Prótons , Água/químicaRESUMO
The fatty acid-based microparticles containing iron oxide nanoparticles and paclitaxel (PAX) are a viable proposition for the treatment of lung cancer. The microparticles inhaled as a dry powder can be guided to selected locations using an external magnetic field, and when accumulated there, the active compound release can be triggered by local hyperthermia. However, this general strategy requires that the active compound is released from microparticles and can reach the targeted cells before microparticles are removed. Isothermal titration calorimetry was used to demonstrate that the components of microparticles were released and transferred to albumins and lipid bilayers. The morphology of the measured particulates was studied with scanning electron microscopy and dynamic light scattering. To determine the cytotoxicity of microparticles, cell culture studies were done. It has been shown that the transfer efficiency depends predominantly on the fatty acid composition of microparticles, which, together with the active ingredient, accumulate predominantly in membrane structures after being released from microparticles and before entering the cytoplasm. The release process is sufficient; hence, paclitaxel-loaded microparticles effectively suppressed the proliferation of A549 human lung epithelial cells of malignant origin (IC50 values for both lauric acid-based and myristic/palmitic-based microparticles containing paclitaxel were below 0.375 µg/mL), while reference microparticles were noncytotoxic.
Assuntos
Ácidos Graxos , Neoplasias Pulmonares , Células A549 , Portadores de Fármacos , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Microscopia Eletrônica de Varredura , Paclitaxel/toxicidade , Tamanho da PartículaRESUMO
Lipid bilayers are active participants in many crucial biological processes. They can be observed in different phases, liquid and solid, respectively. The liquid phase is predominant in biological systems. The solid phase, both crystalline and gel phases, is under investigation due to its resilience to mechanical stress and tight packing of lipids. The mechanical properties of lipids affect their dynamics, therefore influencing the transformation of cell plasma and the endomembrane. Mechanical properties of lipid bilayers are also an important parameter in the design and production of supramolecular lipid-based drug delivery systems. To this end, in this work, we focused on investigating the effect of solid phases of lipid bilayers on their structural parameters and mechanical properties using theoretical molecular dynamics studies on atomistic models of whole vesicles. Those include area per lipid, membrane thickness, density vesicle profiles, bending rigidity coefficient, and area compressibility. Additionally, the bending rigidity coefficient was measured using the flicker noise spectroscopy. The two approaches produced very similar and consistent results. We showed that, contrary to our expectations, bending rigidity coefficients of solid-ordered bilayers for vesicles decreased with an increase in lipid transition temperature. This tendency was reverse in planar systems. Additionally, we have observed an increase of membrane thickness and area compressibility and a decrease of area per lipid. We hope these results will provide valuable mechanical insight for the behavior in solid phases and differences between spherical and planar confirmations.
RESUMO
Drug delivery vectors are widely applied to increase drug efficacy while reducing the side effects and potential toxicity of a drug. They allow for patient-tailored therapy, dose titration, and therapeutic drug monitoring. A major part of drug delivery systems makes use of large nanocarriers: liposomes or virus-like particles (VLPs). These systems allow for a relatively large amount of cargo with good stability of vectors, and they offer multiple options for targeting vectors in vivo. Here we discuss endocytic pathways that are available for drug delivery by large nanocarriers. We focus on molecular aspects of the process, including an overview of potential molecular targets for studies of drug delivery vectors and for future solutions allowing targeted drug delivery.
Assuntos
Preparações de Ação Retardada/química , Endocitose/efeitos dos fármacos , Nanocápsulas/química , Animais , Transporte Biológico , Biomarcadores/metabolismo , Preparações de Ação Retardada/metabolismo , Composição de Medicamentos , Humanos , Lipossomos/química , Terapia de Alvo Molecular , Tamanho da Partícula , Polímeros/químicaRESUMO
Recent interest in the role of ascorbate in crucial metabolic processes is driven by the growing number of medical reports that show beneficial effects of ascorbate supplementation for maintaining general well-being and recovery from a variety of medical conditions. The effect of ascorbate on the local body environment highly depends on its local concentration; at low concentrations it can cause the reduction of reactive oxygen and facilitate activities of enzymes, while at high concentrations it generates free radicals by reducing ferric ions. Ascorbate serving as an electron donor assists the iron-containing proteins and the iron transfer between various aqueous compartments. These functions require effective and adjustable mechanisms responsible for ascorbate biodistribution. In the paper we propose a new biophysical model of ascorbate redistribution between various aqueous body compartments. It combines recent experimental evidence regarding the ability of ascorbate to cross the lipid bilayer by unassisted diffusion, with active transport by well-characterized sodium vitamin C transporter (SVCT) membrane proteins. In the model, the intracellular concentration of ascorbate is maintained by the balance of two opposing fluxes: fast active and slow passive transport. The model provides a mechanistic understanding of ascorbate flux across the epidermal barrier in the gut as well as the role of astrocytes in ascorbate recycling in the brain. In addition, ascorbate passive diffusion across biological membranes, which depends on membrane electric potentials and pH gradients, provides the rationale for the correlation between ascorbate distribution and the transfer of iron ions inside a cell. The proposed approach provides, for the first time, a mechanistic account of processes leading to ascorbate physiological and cellular distribution, which helps to explain numerous experimental and clinical observations.
Assuntos
Ácido Ascórbico/metabolismo , Ácido Ascórbico/farmacocinética , Animais , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Homeostase/efeitos dos fármacos , Humanos , Concentração de Íons de Hidrogênio , Bicamadas Lipídicas , Modelos Biológicos , Transportadores de Sódio Acoplados à Vitamina C/metabolismoRESUMO
Vitamin C is the exogenous compound necessary for a variety of metabolic processes; therefore, the efficient delivery is critical for the maintenance of body homeostasis. Vitamin C pharmacokinetics and low quantities in processed foodstuff, necessitates its continuous supplementation. In the paper, we present the new liposomal formulation of vitamin C free of harmful organic solvents. The formulation was quantitatively characterized with respect to its chemically composition and nano-structuring. The vitamin C accessibility to cells from the formulation was evaluated using evidence derived from experiments performed on cell cultures. Finally, the enhanced bioavailability of vitamin C from the formulation was demonstrated in the medical experiment.
Assuntos
Ácido Ascórbico/administração & dosagem , Ácido Ascórbico/farmacocinética , Administração Oral , Ácido Ascórbico/química , Disponibilidade Biológica , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Composição de Medicamentos , Humanos , LipossomosRESUMO
BACKGROUND: Liposomes serve as delivery systems for biologically active compounds. Existing technologies inefficiently encapsulate large hydrophilic macromolecules, such as PVP-conjugated chlorin e6 (Photolon). This photoactive drug has been widely tested for therapeutic applications, including photodynamic reduction of atherosclerotic plaque. METHODS: A novel formulation of Photolon was produced using "gel hydration technology". Its pharmacokinetics was tested in Sus scrofa f. domestica. Its cellular uptake, cytotoxicity, and ability to induce a phototoxic reaction were demonstrated in J774A.1, RAW264.7 macrophages, and vascular smooth muscle (T/G HA-VSMC) as well as in vascular endothelial (HUVEC) cells. RESULTS: Developed liposomes had an average diameter of 124.7 ± 0.6 nm (polydispersity index (PDI) = 0.055) and contained >80% of Photolon). The half-life of formulation in S. scrofa was 20 min with area under the curve (AUC) equal to 14.7. The formulation was noncytotoxic in vitro and was rapidly (10 min) and efficiently accumulated by macrophages, but not T/G HA-VSMC or HUVEC. The accumulated quantity of photosensitizer was sufficient for induction of phototoxicity in J774A.1, but not in T/G HA-VSMC. CONCLUSIONS: Due to the excellent physical and pharmacokinetic properties and selectivity for macrophages, the novel liposomal formulation of Photolon is a promising therapeutic candidate for use in arteriosclerosis treatment when targeting macrophages but not accompanying vascular tissue is critical for effective and safe therapy.
Assuntos
Lipossomos , Fotoquimioterapia , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Porfirinas/química , Porfirinas/farmacologia , Animais , Linhagem Celular , Clorofilídeos , Composição de Medicamentos , Humanos , Lipossomos/química , Lipossomos/ultraestrutura , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Fotoquimioterapia/métodos , Placa Aterosclerótica/etiologia , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologia , Placa Aterosclerótica/terapia , Espécies Reativas de OxigênioRESUMO
Bending rigidity coefficient describes propensity of a lipid bilayer to deform. In order to measure the parameter experimentally using flickering noise spectroscopy, the microscopic imaging is required, which necessitates the application of giant unilamellar vesicles (GUV) lipid bilayer model. The major difficulty associated with the application of the model is the statistical character of GUV population with respect to their size and the homogeneity of lipid bilayer composition, if a mixture of lipids is used. In the paper, the bending rigidity coefficient was measured using the fluorescence-enhanced flicker-noise spectroscopy. In the paper, the bending rigidity coefficient was determined for large populations of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphocholine vesicles. The quantity of obtained experimental data allows to perform statistical analysis aiming at the identification of the distribution, which is the most appropriate for the calculation of the value of the membrane bending rigidity coefficient. It has been demonstrated that the bending rigidity coefficient is characterized by an asymmetrical distribution, which is well approximated with the gamma distribution. Since there are no biophysical reasons for that we propose to use the difference between normal and gamma fits as a measure of the homogeneity of vesicle population. In addition, the effect of a fluorescent label and types of instrumental setups on determined values has been tested. Obtained results show that the value of the bending rigidity coefficient does not depend on the type of a fluorescent label nor on the type of microscope used.
Assuntos
Fluorescência , Bicamadas Lipídicas/química , Glicerilfosforilcolina/química , Fosfatidilcolinas/química , Espectrometria de Fluorescência , Lipossomas Unilamelares/químicaRESUMO
Octenidine dihydrochloride is an effective antiseptic compound which mode of action is based on destabilization plasma membrane of microorganisms. This ensures that microorganisms cannot develop the drug resistance in a straightforward way, as the entire cellular structure, rather than specific molecular target is affected. Since the octenidine is a hydrophobic compound, it requires organic solvent such as phenoxyethanol in order to be effectively administered. However, the presence of phenoxyethanol has strong irritating effect, particularly when applied on open wounds and mucous membranes. Phospholipids are known as neutral excipients free of side effects and in their aggregated form may serve as solvent for octenidine. In this article, we propose a new antiseptic formulation composed of equimolar ratio of lipids and octenidine. The resulting particles are â¼4 nm in diameter showing that their topology is different from that known for liposomes. The new formulation has proven to be equally effective as octenidine dihydrochloride formulation marketed under the name of Octenisept®. The main advantage of the new formulation is that it does not contain phenoxyethanol, which opens new possibilities for broader application spectrum of octenidine, including treatments of mucous membranes and open wounds.
Assuntos
Anti-Infecciosos Locais/química , Composição de Medicamentos/métodos , Lipídeos/química , Nanopartículas/química , Piridinas/química , Anti-Infecciosos Locais/administração & dosagem , Escherichia coli/efeitos dos fármacos , Etilenoglicóis/química , Corantes Fluorescentes , Iminas , Mucosa/efeitos dos fármacos , Tamanho da Partícula , Piridinas/administração & dosagem , Solventes/química , Propriedades de SuperfícieRESUMO
Lipid bilayer is the main constitutive element of biological membrane, which confines intracellular space. The mechanical properties of biological membranes may be characterized by various parameters including membrane stiffness or membrane bending rigidity, which can be measured using flicker noise spectroscopy. The flicker noise spectroscopy exploits the spontaneous thermal undulations of the membrane. The method is based on the quantitative analysis of a series of microscopic images captured during thermal membrane fluctuations. Thus, measured bending rigidity coefficient depends on the image quality as well as the selection of computational tools for image processing and mathematical model used. In this work scanning and spinning disc confocal microscopies were used to visualize fluctuating membranes of giant unilamellar vesicles. The bending rigidity coefficient was calculated for different acquisition modes, using different fluorescent probes and different image processing methods. It was shown that both imaging approaches gave similar bending coefficient values regardless of acquisition time. Using the developed methodology the effect of fluorescent probe type and aqueous phase composition on the value of the membrane bending rigidity coefficient was measured. Specifically it was found that the bending rigidity coefficient of DOPC bilayer in water is smaller than that determined for POPC membrane. It has been found that the POPC and DOPC bending rigidities coefficient in sucrose solution was lower than that in water. Fluorescence imaging makes possible the quantitative analysis of membrane mechanical properties of inhomogeneous membrane.
Assuntos
Corantes Fluorescentes/química , Bicamadas Lipídicas/química , Fosfatidilcolinas/química , Espectrometria de FluorescênciaRESUMO
Organotin compounds, being biologically active, affect a variety of cellular functions due to their ability to accumulate in and penetrate biological membranes. These compounds influence the distribution of electrostatic charges, alter organization, disrupt molecular dynamics and change mechanical properties of biological membranes. It was found that the membrane/water partition coefficient equals 4, a value significantly higher than octanol/water partition coefficient. In addition, the effect of di- and tri-phenyltin chlorides on the mechanics of model lipid membranes was measured for the first time. It has been determined that phenyltins affect the global model lipid bilayer properties by reducing the membrane expansion modulus, when measured using micromanipulation technique, and elevating the bending rigidity coefficient of the lipid bilayer, as determined with the flickering noise spectroscopy. In addition, the elevated water permeability shows that phenyltins also cause the local defects formation in the lipid bilayer, i.e. lipid pores. These data shows that phenyltins may interfere indirectly with variety cellular processes by altering non-specifically the entire cellular membrane system. Accordingly, when phenyltins are added to macrophages in culture, they inflict massive alterations of cell morphology and interfere with membrane-associated processes, as visualized using fluorescence labelling of selected subcellular compartments.
Assuntos
Bicamadas Lipídicas/química , Macrófagos/efeitos dos fármacos , Compostos Orgânicos de Estanho/farmacologia , Fosfatidilcolinas/química , Lipossomas Unilamelares/química , Laranja de Acridina/metabolismo , Animais , Linhagem Celular , Cloretos/química , Relação Dose-Resposta a Droga , Peróxido de Hidrogênio/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Permeabilidade/efeitos dos fármacos , Água/metabolismoRESUMO
The ionic composition of intracellular space is rigorously maintained in the expense of high-energy expenditure. It has been recently postulated that the cytoplasmic ionic composition is optimized so the energy cost of the fluctuations of calcium ion concentration is minimized. Specifically, thermodynamic arguments have been produced to show that the presence of potassium ions at concentrations higher than 100 mM reduce extend of the energy dissipation required for the dilution of calcium cations. No such effect has been measured when sodium ions were present in the solution or when the other divalent cation magnesium was diluted. The experimental observation has been interpreted as the indication of the formation of ionic clusters composed of calcium, chloride and potassium. In order to test the possibility that such clusters may be preserved in biological space, the thermodynamics of ionic mixtures dilution in solutions containing albumins and model lipid bilayers have been measured. Obtained thermograms clearly demonstrate that the energetics of calcium/potassium mixture is qualitatively different from calcium/sodium mixture indicating that the presence of the biologically relevant quantities of proteins and membrane hydrophilic surfaces do not interfere with the properties of the intracellular aqueous phase.
Assuntos
Biopolímeros/química , Cátions/química , Misturas Complexas/química , Modelos Químicos , Soluções/química , Termodinâmica , Simulação por Computador , Técnicas de Diluição do IndicadorRESUMO
Fluorescence Correlation Spectroscopy (FCS) is a technique, which allows determination of the diffusion coefficient and concentration of fluorescent objects suspended in the solution. The measured parameter is the fluctuation of the fluorescence signal emitted by diffusing molecules. When 100 nm DOPC vesicles labeled with various fluorescent dyes (Fluorescein-PE, NBD-PE, Atto488 DOPE or ßBodipy FL) were measured, different values of diffusion coefficients have been obtained. These diffusion coefficients were different from the expected values measured using the dynamic light scattering method (DLS). The FCS was initially developed for solutions containing small fluorescent molecules therefore the observed inconsistency may result from the nature of vesicle suspension itself. The duration of the fluorescence signal may depend on the following factors: the exposure time of the labeled object to the excitation beam, the photo-physical properties (e.g., stability) of a fluorophore, the theoretical model used for the calculations of the diffusion coefficient and optical properties of the vesicle suspension. The diffusion coefficients determined for differently labeled liposomes show that its dependence on vesicle size and quantity of fluorescent probed used for labeling was significant demonstrating that the fluorescence properties of the fluorophore itself (bleaching and/or blinking) were critical factors for a correct outcome of FCS experiment. The new, based on combined FCS and DLS measurements, method for the determination of the focal volume prove itself to be useful for the evaluation of a fluorescence dye with respect to its applicability for FCS experiment.
Assuntos
Fluorescência , Corantes Fluorescentes/química , Glicerilfosforilcolina/análogos & derivados , Lipossomos/química , Espectrometria de Fluorescência/métodos , Difusão , Glicerilfosforilcolina/química , Modelos Teóricos , FosfatidilcolinasRESUMO
Catestatin, a cationic and hydrophobic 21-amino acid fragment of chromogranin A, is known to be a non-competitive nicotinic antagonist acting through nicotinic acetylcholine receptors (nAChRs) to inhibit catecholamine release. Since this receptor is the target of several neuronal and non-neuronal disorder prophylaxes and treatments, this study aims at the elucidation of the binding of human catestatin to the entire nAChR reconstructed in lipid bilayers by means of docking followed by full atomistic molecular dynamics simulations. The obtained results show that the minimum free energy for the binding of the peptide and the receptor attains minimal values for locations at the pore site and in the outer beta subunit. This result is consistent with previous studies showing that catestatin occludes the pore opening. A new finding is an additional even stronger binding seat at the beta subunit and that membrane presence could be an important factor. Specific amino acids involved in catestatin binding have been identified, indicating targets for point mutation studies. In addition to improving the understanding of the interaction between the peptide and muscle-type and even other nAChR subtypes, the results of this study provide directions for future peptidomimetic research.
Assuntos
Aminoácidos/análise , Aminoácidos/química , Cromogranina A/química , Simulação de Dinâmica Molecular , Fragmentos de Peptídeos/química , Receptores Nicotínicos/química , Sítios de Ligação , HumanosRESUMO
Biological systems are characterized by directional and precisely controlled flow of matter and information along with the maintenance of their structural patterns. This is possible thanks to sequential transformations of information, energy and structure carried out by molecular machines. The new perception of biological systems, including their mechanical aspects, requires the implementation of tools and approaches previously developed for engineering sciences. In this review paper, a biological system is presented in a new perspective as an ensemble of coordinated molecular devices functioning in the limited space confined by the biological membrane. The working of a molecular machine is presented using the example of F0F1 ATPase, and the general conditions necessary for the coordination of a large number of functional units are described.
Assuntos
ATPases Translocadoras de Prótons/química , Transporte Biológico Ativo , ATPases Translocadoras de Prótons/metabolismoRESUMO
The changing environment and modified lifestyles have meant that many vitamins and minerals are deficient in a significant portion of the human population. Therefore, supplementation is a viable nutritional approach, which helps to maintain health and well-being. The supplementation efficiency of a highly hydrophobic compound such as cholecalciferol (logP > 7) depends predominantly on the formulation. To overcome difficulties associated with the evaluation of pharmacokinetics of cholecalciferol, a method based on the short time absorption data in the clinical study and physiologically based mathematical modeling is proposed. The method was used to compare pharmacokinetics of liposomal and oily formulations of vitamin D3. The liposomal formulation was more effective in elevating calcidiol concentration in serum. The determined AUC value for liposomal vitamin D3 formulation was four times bigger than that for the oily formulation.
RESUMO
The effect of lipid oxidation on water permeability of phosphatidylcholine membranes was investigated by means of both scattering stopped flow experiments and atomistic molecular dynamics simulations. Formation of water pores followed by a significant enhancement of water permeability was observed. The molecules of oxidized phospholipids facilitate pore formation and subsequently stabilize water in the membrane interior. A wide range of oxidation ratios, from 15 to 100 mol%, was considered. The degree of oxidation was found to strongly influence the time needed for the opening of a pore. In simulations, the oxidation ratio of 75 mol% was found to be a threshold for spontaneous pore formation in the tens of nanosecond timescale, whereas 15 mol% of oxidation led to significant water permeation in the timescale of seconds. Once a pore was formed, the water permeability was found to be virtually independent of the oxidation ratio.
Assuntos
Bicamadas Lipídicas/química , Fosfatidilcolinas/química , Água/química , Simulação de Dinâmica Molecular , Oxirredução , PermeabilidadeRESUMO
Ascorbate is an important element of a variety of cellular processes including the control of reactive oxygen species levels. Since reactive oxygen species are implicated as a key factor in tumorigenesis and antitumor therapy, the injection of a large amount of ascorbate is considered beneficial in cancer therapy. Recent studies have shown that ascorbate can cross the plasma membrane through passive diffusion. In contrast to absorption by active transport, which is facilitated by transport proteins (SVCT1 and SVCT2). The passive diffusion of a weak acid across membranes depends on the electrostatic potential and the pH gradients. This has been used to construct a new theoretical model capable of providing steady-state ascorbate concentration in the intracellular space and evaluating the time needed to reach it. The main conclusion of the analysis is that the steady-state intracellular ascorbate concentration weakly depends on its serum concentration but requires days of exposure to saturate. Based on these findings, it can be hypothesized that extended oral ascorbate delivery is possibly more effective than a short intravenous infusion of high ascorbate quantities.
Assuntos
Ácido Ascórbico/metabolismo , Espaço Intracelular/metabolismo , Potenciais da Membrana/fisiologia , Neoplasias/terapia , Linhagem Celular Tumoral , Espaço Extracelular/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Modelos Biológicos , Análise Numérica Assistida por Computador , Transportadores de Sódio Acoplados à Vitamina C/metabolismo , Fatores de TempoRESUMO
Extrusion is a popular method for producing homogenous population of unilamellar liposomes. The technique relies on forcing a lipid suspension through cylindrical pores in a polycarbonate membrane. The quantification of the extrusion and/or recalibration processes make possible the acquisition of experimental data, which can be correlated with the mechanical properties of the lipid bilayer. In this work, the force needed for the extrusion process was correlated with the mechanical properties of a lipid bilayer derived from other experiments. Measurements were performed using a home-made dedicated device capable of maintaining a stable volumetric flux of a liposome suspension through well-defined pores and to continuously measure the extrusion force. Based on the obtained results, the correlation between the lipid bilayer bending rigidity and extrusion force was derived. Specifically, it was found that the bending rigidity of liposomes formed from well-defined lipid mixtures agrees with data obtained by others using flicker-noise spectroscopy or micromanipulation. The other issue addressed in the presented studies was the identification of molecular mechanisms leading to the formation of unilamellar vesicles in the extrusion process. Finally, it was demonstrated that during the extrusion, lipids are not exchanged between vesicles, i.e., vesicles can divide but no membrane fusion or lipid exchange between bilayers was detected.