Browsing by Subject "Liposomes"
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Item Phase-separated liposomes for efficient macromolecular delivery(2018-07-25) Imam, Zachary Ibrahim; Stachowiak, Jeanne Casstevens; Smyth, Hugh; Baker, Aaron; Gordon, Vernita; Zoldan, JanetaFrom viruses to organelles, fusion of biological membranes is used by diverse biological systems to deliver molecules across membrane barriers. Membrane fusion is a potentially efficient mechanism for the delivery of macromolecular therapeutics and chemotherapeutics to the cellular cytoplasm. However, a key shortcoming of existing fusogenic delivery systems is their inefficiency, requiring a high concentration of fusion-promoting lipids to cross cellular membrane barriers. To address this limitation, my work has explored the extent to which membrane fusion can be amplified through the process of lipid membrane phase-separation to concentrate fusion-promoting lipids within distinct regions on the membrane surface. Towards building a fusogenic phase-separated liposomal system for direct delivery, I began by investigating the impact of incorporating membrane-bound polymers onto the surface of phase-separated liposomes. Membrane-bound polymers like polyethylene glycol (PEG) are used to increase the circulation time of liposomes in vivo and are known to influence lipid phase behavior. My work demonstrates that membrane-bound PEGs crowded on liposome surfaces can generate significant steric pressure, which is sufficient to destabilize phase-separated lipid domains. These data show the importance of optimizing membrane coverage of polymers on the surface of phase-separated liposomes. I next report the development of fusogenic phase-separated liposomes. Specifically, I show that concentrating fusion-promoting lipids within phase-separated domains on liposome surfaces significantly increases the efficiency of liposome fusion with model membranes and cellular membranes. In particular, membrane phase-separation enhances delivery of lipids and model macromolecules to the cytoplasm by at least 4-fold relative to homogenous liposomes. Furthermore, I demonstrate that phase-separated fusogenic liposomes can be loaded with the chemotherapeutic doxorubicin using conventional active loading protocols. These fusogenic phase-separated liposomes reduced the therapeutically effective dose of encapsulated doxorubicin by 4-fold relative to homogeneous liposomes and control liposomes lacking fusogenic lipids. My findings demonstrate that membrane phase-separation can enhance membrane fusion by locally concentrating fusion-promoting lipids on the surface of liposomes. This work represents the first application of lipid membrane phase-separation in the design of biomaterials-based delivery systems. Additionally, these results lay the ground work for developing fusogenic liposomes that are triggered by physical and molecular cues associated with target cellsItem Sorption of selected endocrine disrupters by synthetic membrane vesicles and effects of natural organic matter(2002-05) Yamamoto, Hiroshi, 1973-; Liljestrand, Howard M. (Howard Michael)In this study, synthetic membrane vesicles (liposomes) were prepared from phospholipids and cholesterol as a model biological phase, and the bioavailability of four estrogens and five estrogenic compounds were evaluated by determining the sorption coefficients (Klipw values) into liposomes. Since natural organic matter (NOM) is likely to bind hydrophobic organics such as endocrine disrupters and influence their bioavailability and the fate in the environment, the effects of a wide variety of model NOM compounds, including humic substances, polysaccharides, and tannic acid, on the liposome-water system were investigated by determining the sorption coefficients onto NOM (Koc values). Reasonably reproducible preparations of liposome suspensions were characterized by quasi-elastic laser light scattering (QELS) analysis and transmission electron microscopy (TEM). Reproducible measurements of Klipw values were obtained by the equilibrium dialysis technique. The Klipw values determined strongly depend on the lipid components of the liposome. Liposome with shorter acyl-chain or increased number of double bonds had higher values of Klipw, while those including cholesterol had significantly lower Klipw values than those without. Moreover, log Klipw values of the selected endocrine disrupters did not show a strong linear correlation with their log Kow. Thus, the use of linear free energy relationships (LFER) to estimate Klipw values is limited for these compounds with a wide variety of chemical structures. Koc values for the selected NOM compounds were determined by the fluorescence quenching technique. The Koc values for the selected compounds had a good linear correlation with the absorptivity at 272 nm and the concentration of phenolic groups of each NOM. However, log Koc showed no correlation with log Kow for the selected endocrine disrupters. Nominal sorption coefficients into liposome ( ) slightly decreased with an increase of NOM concentration, but the decrease was not significant at the highest NOM concentration. Simple models developed in this study showed satisfactory agreement of the estimated K values with those measured for two humic acids. Other linear sorption models were developed for activated sludge treatment and found to agree with literature results. Simple models were also developed to estimate the relative health-related risks of endocrine disrupters.