Browsing by Subject "LIPOSOMES"

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  • Costa, Clarinda; Liu, Zehua; Martins, João Pedro; Correia, Alexandra; Rahikkala, Antti Tuomas Antero; Li, Wei; Seitsonen, Jani; Ruokolainen, Janne; Hirvonen, Sami-Pekka; Aguiar- Ricardo, Ana; Corvo, M. Luísa; Santos, Hélder A. (2020)
    Here, a continuous two-step glass-capillary microfluidic technique to produce a multistage oral delivery system is reported. Insulin is successfully encapsulated into liposomes, which are coated with chitosan to improve their mucoadhesion. The encapsulation in an enteric polymer offers protection from the harsh gastric conditions. Insulin permeability is enhanced across an intestinal monolayer.
  • Allolio, Christoph; Magarkar, Aniket; Jurkiewicz, Piotr; Baxova, Katarina; Javanainen, Matti; Mason, Philip E.; Sachl, Radek; Cebecauer, Marek; Hof, Martin; Horinek, Dominik; Heinz, Veronika; Rachel, Reinhard; Ziegler, Christine M.; Schröfel, Adam; Jungwirth, Pavel (2018)
    Arginine-rich cell-penetrating peptides do not enter cells by directly passing through a lipid membrane; they instead passively enter vesicles and live cells by inducing membrane multilamellarity and fusion. The molecular picture of this penetration mode, which differs qualitatively from the previously proposed direct mechanism, is provided by molecular dynamics simulations. The kinetics of vesicle agglomeration and fusion by an iconic cell-penetrating peptide-nonaarginine-are documented via real-time fluorescence techniques, while the induction of multilamellar phases in vesicles and live cells is demonstrated by a combination of electron and fluorescence microscopies. This concert of experiments and simulations reveals that the identified passive cell penetration mechanism bears analogy to vesicle fusion induced by calcium ions, indicating that the two processes may share a common mechanistic origin.
  • Balasubramanian, Vimalkumar; Poillucci, Andrea; Correia, Alexandra; Zhang, Hongbo; Celia, Christian; Santos, Helder A. (2018)
    Organelles of eukaryotic cells are structures made up of membranes, which carry out a majority of functions necessary for the surviving of the cell itself. Organelles also differentiate the prokaryotic and eukaryotic cells, and are arranged to form different compartments guaranteeing the activities for which eukaryotic cells are programmed. Cell membranes, containing organelles, are isolated from cancer cells and erythrocytes and used to form biocompatible and long circulating ghost nanoparticles delivering payloads or catalyzing enzymatic reactions as nanoreactors. In this attempt, red blood cell membranes were isolated from erythrocytes, and engineered to form nanoerythrosomes (NERs) of 150 nm. The horseradish peroxidase, used as an enzyme model, was loaded inside the aqueous compartment of NERs, and its catalytic reaction with Resorufm was monitored. The resulting nanoreactor protected the enzyme from proteolytic degradation, and potentiated the enzymatic reaction in situ as demonstrated by maximal velocity (V-max) and Michaelis constant (K-m), thus suggesting the high catalytic activity of nanoreactors compared to the pure enzymes.
  • Magarkar, Aniket; Dhawan, Vivek; Kallinteri, Paraskevi; Viitala, Tapani; Elmowafy, Mohammed; Rog, Tomasz; Bunker, Alex (2014)
  • Poojari, Chetan; Zak, Agata; Dzieciuch-Rojek, Monika; Bunker, Alex; Kepczynski, Mariusz; Rog, Tomasz (2020)
    Cholesterol plays a crucial role in modulating the physicochemical properties of biomembranes, both increasing mechanical strength and decreasing permeability. Cholesterol is also a common component of vesicle-based delivery systems, including liposome-based drug delivery systems (LDSs). However, its effect on the partitioning of drug molecules to lipid membranes is very poorly recognized. Herein, we performed a combined experimental/computational study of the potential for the use of the LDS formulation for the delivery of the antifungal drug itraconazole (ITZ). We consider the addition of cholesterol to the lipid membrane. Since ITZ is only weakly soluble in water, its bioavailability is limited. Use of an LDS has thus been proposed. We studied lipid membranes composed of cholesterol, 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine (POPC), and ITZ using a combination of computational molecular dynamics (MD) simulations of lipid bilayers and Brewster angle microscopy (BAM) experiments of monolayers. Both experimental and computational results show separation of cholesterol and ITZ. Cholesterol has a strong preference to orient parallel to the bilayer normal. However, ITZ, a long and relatively rigid molecule with weakly hydrophilic groups along the backbone, predominantly locates below the interface between the hydrocarbon chain region and the polar region of the membrane, with its backbone oriented parallel to the membrane surface; the orthogonal orientation in the membrane could be the cause of the observed separation. In addition, fluorescence measurements demonstrated that the affinity of ITZ for the lipid membrane is decreased by the presence of cholesterol, which is thus probably not a suitable formulation component of an LDS designed for ITZ delivery.
  • Kasparyan, Gari; Poojari, Chetan; Rog, Tomasz; Hub, Jochen S. (2020)
    Itraconazole is a triazole drug widely used in the treatment of fungal infections, and it is in clinical trials for treatment of several cancers. However, the drug suffers from poor solubility, while experiments have shown that itraconazole delivery in liposome nanocarriers improves both circulation half-life and tissue distribution. The drug release mechanism from the nanocarrier is still unknown, and it depends on several factors including membrane stability against defect formation. In this work, we used molecular dynamics simulations and potential of mean force (PMF) calculations to quantify the influence of itraconazole on pore formation over lipid membranes, and we compared the effect by itraconazole with a pore-stabilizing effect by the organic solvent dimethyl sulfoxide (DMSO). According to the PMFs, both itraconazole and DMSO greatly reduce the free energy of pore formation, by up to similar to 20 kJ mol(-1). However, whereas large concentrations of itraconazole of 8 mol % (relative to lipid) were required, only small concentrations of a few mole % DMSO (relative to water) were sufficient to stabilize pores. In addition, itraconazole and DMSO facilitate pore formation by different mechanisms. Whereas itraconazole predominantly aids the formation of a partial defect with a locally thinned membrane, DMSO mainly stabilizes a transmembrane water needle by shielding it from the hydrophobic core. Notably, the two distinct mechanisms act cooperatively upon adding both itraconazole and DMSO to the membrane, as revealed by an additional reduction of the pore free energy. Overall, our simulations reveal molecular mechanisms and free energies of membrane pore formation by small molecules. We suggest that the stabilization of a locally thinned membrane as well as the shielding of a transmembrane water needle from the hydrophobic membrane core may be a general mechanism by which amphiphilic molecules facilitate pore formation over lipid membranes at sufficient concentrations.
  • Wilkosz, Natalia; Rissanen, Sami; Cyza, Malgorzata; Szybka, Renata; Nowakowska, Maria; Bunker, Alex; Rog, Tomasz; Kepczynski, Mariusz (2017)
    Uptake of piroxicam, a non-steroidal anti-inflammatory drug, from the intestines after oral intake is limited due to its low solubility and its wide use is associated with several side effects related to the gastrointestinal tract. In this study, all-atom molecular dynamics (MD) simulations and fluorescent spectroscopy were employed to investigate the interaction of piroxicam in neutral, zwitterionic, and cationic forms with lipid bilayers composed of phosphatidylcholine, cholesterol, and PEGylated lipids. Our study was aimed to assess the potential for encapsulation of piroxicam in liposomal carriers and to shed more light on the process of gastrointestinal tract injury by the drug. Through both the MD simulations and laser scanning confocal microscopy, we have demonstrated that all forms of piroxicam can associate with the lipid bilayers and locate close to the water-membrane interface. Conventional liposomes used in drug delivery are usually stabilized by the addition of cholesterol and have their bloodstream lifetime extended through the inclusion of PEGylated lipids in the formulation to create a protective polymer corona. For this reason, we tested the effect of these two modifications on the behavior of piroxicam in the membrane. When the bilayer was PEGylated, piroxicam localize to the PEG layer and within the lipid headgroup region. This suggests that PEGylated liposomes are capable of carrying a larger quantity of piroxicam than the conventional ones. (C) 2017 Elsevier B.V. All rights reserved.
  • Ridolfo, Roxane; Tavakoli, Shirin; Junnuthula, Vijayabhaskarreddy; Williams, David S.; Urtti, Arto; van Hest, Jan C. M. (2021)
    Nanoparticle morphology (size, shape, and composition) and surface chemistry are the determining factors underpinning the efficacy of such materials in therapeutic applications. The size, shape, and surface chemistry of a nanoparticle can strongly influence key properties such as interactions with diverse biological fluids and interfaces and, in turn, impact the delivery of bioactive cargo, modulating therapeutic performance. This is exemplified in ocular drug delivery, where potential therapeutics must navigate complex biological media such as the gel-like vitreal fluid and the retina. Biodegradable block copolymer amphiphiles are a robust tool for the engineering of various types of self-assembled nanoparticles with diverse morphologies ranging from spherical and tubular polymersomes to spherical and worm-like micelles. Here, we explore the effect of morphological features such as shape and surface chemistry upon the interactions of a series of copolymer nanoparticles with retinal (ARPE-19) cells and the release of a low solubility drug (dexamethasone) that is currently used in ocular therapy and study their diffusion in vitreous using ex vivo eyes. We demonstrate that both aspect ratio and surface chemistry of nanoparticles will influence their performance in terms of cell uptake, drug release, and diffusion with high aspect ratio shapes demonstrating enhanced properties in relation to their spherical counterparts.
  • Marwah, Megha; Magarkar, Aniket; Ray, Debes; Aswal, Vinod; Bunker, Alex; Nagarsenker, Mangal (2018)
    Glyceryl monostearate (GMS) is a single-tailed lipidic monoglyceride commonly used as a nontoxic food additive. In this study, we have investigated GMS, specifically its self-assembling properties and subsequent application in drug delivery. Results from in silico modeling, corroborated by complementary small-angle neutron scattering, demonstrated vesicle formation; associated phase transitions were analyzed using differential scanning calorimetry; dynamic light scattering revealed particle size alterations that occurred in the transition region. Spherical morphology of unilamellar vesicles was visualized using transmission electron microscopy imaging. Further, hydrophilic and hydrophobic drug loading in GMS vesicles and their amenability to surface modification for hepatic targeting have, in this study, been both predicted through molecular simulation study and demonstrated experimentally. The influence of hepatotropic ligands on the stability of drug-loaded GMS vesicles vis-a-vis cholesterol has also been investigated; the resulting GMS-based drug delivery vehicle, its properties enhanced through surface decoration, is envisaged to achieve targeted delivery of its payload to hepatocytes.
  • Dusa, Filip; Chen, Wen; Witos, Joanna; Rantamäki, Antti; King, Alistair; Sklavounos, Evangelos; Roth, Michal; Wiedmer, Susanne (2020)
    The cell membrane is mainly composed of lipid bilayers with inserted proteins and carbohydrates. Lipid bilayers made of purified or synthetic lipids are widely used for estimating the effect of target compounds on cell membranes. However, the composition of such biomimetic membranes is much simpler than the composition of biological membranes. Interactions between compounds and simple composition biomimetic membranes might not demonstrate the effect of target compounds as precisely as membranes with compositions close to real organisms. Therefore, the aim of our study is to construct biomimetic membrane closely mimicking the state of natural membranes. Liposomes were prepared from lipids extracted from L-alpha-phosphatidylcholine, Escherichia coli, yeast (Saccharomyces cerevisiae) and bovine liver cells through agitation and sonication. They were immobilized onto silicon dioxide (SiO2) sensor surfaces using N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid buffer with calcium chloride. The biomimetic membranes were successfully immobilized onto the SiO2 sensor surface and detected by nanoplasmonic sensing. The immobilized membranes were exposed to choline carboxylates. The membrane disruption effect was, as expected, more pronounced with increasing carbohydrate chain length of the carboxylates. The results correlated with the toxicity values determined using Vibrio fischeri bacteria. The yeast extracted lipid membranes had the strongest response to introduction of choline laurate while the bovine liver lipid extracted liposomes were the most sensitive towards the shorter choline carboxylates. This implies that the composition of the cell membrane plays a crucial role upon interaction with choline carboxylates, and underlines the necessity of testing membrane systems of different origin to obtain an overall image of such interactions.
  • Kontturi, Leena-Stiina; van den Dikkenberg, Joep; Urtti, Arto; Hennink, Wim E.; Mastrobattista, Enrico (2019)
    The major challenge in the therapeutic applicability of oligonucleotide-based drugs is the development of efficient and safe delivery systems. The carriers should be non-toxic and stable in vivo, but interact with the target cells and release the loaded oligonucleotides intracellularly. We approached this challenge by developing a light-triggered liposomal delivery system for oligonucleotides based on a non-cationic and thermosensitive liposome with indocyanine green (ICG) as photosensitizer. The liposomes had efficient release properties, as 90% of the encapsulated oligonucleotides were released after 1-minute light exposure. Cell studies using an enhanced green fluorescent protein (EGFP)-based splicing assay with HeLa cells showed light-activated transfection with up to 70%-80% efficacy. Moreover, free ICG and oligonucleotides in solution transfected cells upon light induction with similar efficacy as the liposomal system. The light-triggered delivery induced moderate cytotoxicity (25%-35% reduction in cell viability) 1-2 days after transfection, but the cell growth returned to control levels in 4 days. In conclusion, the ICG-based light-triggered delivery is a promising method for oligonucleotides, and it can be used as a platform for further optimization and development.
  • Medina, Tuula Penate; Gerle, Mirko; Humbert, Jana; Chu, Hanwen; Koepnick, Anna-Lena; Barkmann, Reinhard; Garamus, Vasil M.; Sanz, Beatriz; Purcz, Nicolai; Will, Olga; Appold, Lia; Damm, Timo; Suojanen, Juho; Arnold, Philipp; Lucius, Ralph; Willumeit-Roemer, Regina; Acil, Yahya; Wiltfang, Joerg; Goya, Gerardo F.; Glueer, Claus C.; Medina, Oula Penate (2020)
    Simple Summary A novel active release system magnetic sphingomyelin-containing liposome encapsulated with indocyanine green, fluorescent marker, or the anticancer drug cisplatin was evaluated. The liposomal sphingomyelin is a target for the sphingomyelinase enzyme, which is released by stressed cells. Thus, sphingomyelin containing liposomes behave as a sensitizer for biological stress situations. In addition, the liposomes were engineered by adding paramagnetic beads to act as a receiver of outside given magnetic energy. The enzymatic activity towards liposomes and destruction caused by the applied magnetic field caused the release of the content from the liposomes. By using these novel liposomes, we could improve the drug release feature of liposomes. The improved targeting and drug-release were shown in vitro and the orthotopic tongue cancer model in mice optical imaging. The increased delivery of cisplatin prolonged the survival of the targeted delivery group versus free cisplatin. Most available cancer chemotherapies are based on systemically administered small organic molecules, and only a tiny fraction of the drug reaches the disease site. The approach causes significant side effects and limits the outcome of the therapy. Targeted drug delivery provides an alternative to improve the situation. However, due to the poor release characteristics of the delivery systems, limitations remain. This report presents a new approach to address the challenges using two fundamentally different mechanisms to trigger the release from the liposomal carrier. We use an endogenous disease marker, an enzyme, combined with an externally applied magnetic field, to open the delivery system at the correct time only in the disease site. This site-activated release system is a novel two-switch nanomachine that can be regulated by a cell stress-induced enzyme at the cellular level and be remotely controlled using an applied magnetic field. We tested the concept using sphingomyelin-containing liposomes encapsulated with indocyanine green, fluorescent marker, or the anticancer drug cisplatin. We engineered the liposomes by adding paramagnetic beads to act as a receiver of outside magnetic energy. The developed multifunctional liposomes were characterized in vitro in leakage studies and cell internalization studies. The release system was further studied in vivo in imaging and therapy trials using a squamous cell carcinoma tumor in the mouse as a disease model. In vitro studies showed an increased release of loaded material when stress-related enzyme and magnetic field was applied to the carrier liposomes. The theranostic liposomes were found in tumors, and the improved therapeutic effect was shown in the survival studies.
  • Herranz- Blanco, Bárbara; Arriaga, Laura R.; Mäkilä, Ermei; Correia, Alexandra; Shrestha, Neha; Mirza, Sabiruddin; Weitz, David A.; Salonen, Jarno; Hirvonen, Jouni; Santos, Helder A. (2014)
    A reliable microfluidic platform for the generation of stable and monodisperse multistage drug delivery systems is reported. A glass-capillary flow-focusing droplet generation device was used to encapsulate thermally hydrocarbonized porous silicon (PSi) microparticles into the aqueous cores of double emulsion drops, yielding the formation of a multistage PSi–lipid vesicle. This composite system enables a large loading capacity for hydrophobic drugs.
  • Rissanen, Sami; Grzybek, Michal; Orlowski, Adam; Rog, Tomasz; Cramariuc, Oana; Levental, Ilya; Eggeling, Christian; Sezgin, Erdinc; Vattulainen, Ilpo (2017)
    Driven by interactions between lipids and proteins, biological membranes display lateral heterogeneity that manifests itself in a mosaic of liquid-ordered (Lo) or raft, and liquid-disordered (Ld) or non-raft domains with a wide range of different properties and compositions. In giant plasma membrane vesicles and giant unilamellar vesicles, specific binding of Cholera Toxin (CTxB) to GM1 glycolipids is a commonly used strategy to label raft domains or Lo membrane environments. However, these studies often use acyl-chain labeled bodipy-GM1 (bdGM1), whose headgroup accessibility and membrane order or phase partitioning may differ from those of GM1, rendering the interpretation of CTxB binding data quite problematic. To unravel the molecular basis of CTxB binding to GM1 and bdGM1, we explored the partitioning and the headgroup presentation of these gangliosides in the Lo and Ld phases using atomistic molecular dynamics simulations complemented by CTxB binding experiments. The conformation of both GM1 and bdGM1 was shown to be largely similar in the Lo and Ld phases. However, bdGM1 showed reduction in receptor availability when reconstituted into synthetic bilayer mixtures, highlighting that membrane phase partitioning of the gangliosides plays a considerable role in CTxB binding. Our results suggest that the CTxB binding is predominately modulated by the partitioning of the receptor to an appropriate membrane phase. Further, given that the Lo and Ld partitioning of bdGM1 differs from those of GM1, usage of bdGM1 for studying GM1 behavior in cells can lead to invalid interpretation of experimental data.
  • Cheng, Ruoyu; Fontana, Flavia; Xiao, Junyuan; Liu, Zehua; Figueiredo, Patricia; Shahbazi, Mohammad-Ali; Wang, Shiqi; Jin, Jing; Torrieri, Giulia; Hirvonen, Jouni T.; Zhang, Hongbo; Chen, Tongtong; Cui, Wenguo; Lu, Yong; Santos, Helder A. (2020)
    Recently, there has been an increasing interest for utilizing the host immune system to fight against cancer. Moreover, cancer vaccines, which can stimulate the host immune system to respond to cancer in the long term, are being investigated as a promising approach to induce tumor-specific immunity. In this work, we prepared an effective cancer vaccine (denoted as vacosome) by reconstructing the cancer cell membrane, monophosphoryl lipid A as a toll-like receptor 4 agonist, and egg phosphatidylcholine. The vacosome triggered and enhanced bone marrow dendritic cell maturation as well as stimulated the antitumor response against breast cancer 4T1 cells in vitro. Furthermore, an immune memory was established in BALB/c mice after three-time preimmunization with the vacosome. After that, the immunized mice showed inhibited tumor growth and prolonged survival period (longer than 50 days). Overall, our results demonstrate that the vacosome can be a potential candidate for clinical translation as a cancer vaccine.
  • Olzynska, Agnieszka; Kulig, Waldemar; Mikkolainen, Heikki; Czerniak, Tomasz; Jurkiewicz, Piotr; Cwiklik, Lukasz; Rog, Tomasz; Hof, Martin; Jungwirth, Pavel; Vattulainen, Ilpo (2020)
    Cholesterol renders mammalian cell membranes more compact by reducing the amount of voids in the membrane structure. Because of this, cholesterol is known to regulate the ability of cell membranes to prevent the permeation of water and water-soluble molecules through the membranes. Meanwhile, it is also known that even seemingly tiny modifications in the chemical structure of cholesterol can lead to notable changes in membrane properties. The question is, how significantly do these small changes in cholesterol structure affect the permeability barrier function of cell membranes? In this work, we applied fluorescence methods as well as atomistic molecular dynamics simulations to characterize changes in lipid membrane permeability induced by cholesterol oxidation. The studied 7 beta-hydroxycholesterol (7 beta-OH-chol) and 27-hydroxycholesterol (27-OH-chol) represent two distinct groups of oxysterols, namely, ring- and tail-oxidized cholesterols, respectively. Our previous research showed that the oxidation of the cholesterol tail has only a marginal effect on the structure of a lipid bilayer; however, oxidation was found to disturb membrane dynamics by introducing a mechanism that allows sterol molecules to move rapidly back and forth across the membranebobbing. Herein, we show that bobbing of 27-OH-chol accelerates fluorescence quenching of NBD-lipid probes in the inner leaflet of liposomes by dithionite added to the liposomal suspension. Systematic experiments using fluorescence quenching spectroscopy and microscopy led to the conclusion that the presence of 27-OH-chol increases membrane permeability to the dithionite anion. Atomistic molecular dynamics simulations demonstrated that 27-OH-chol also facilitates water transport across the membrane. The results support the view that oxysterol bobbing gives rise to successive perturbations to the hydrophobic core of the membrane, and these perturbations promote the permeation of water and small water-soluble molecules through a lipid bilayer. The observed impairment of permeability can have important consequences for eukaryotic organisms. The effects described for 27-OH-chol were not observed for 7 beta-OH-chol which represents ring-oxidized sterols.