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Liposomes
Published in Danilo D. Lasic, LIPOSOMES in GENE DELIVERY, 2019
Other methods for liposome preparation are detergent depletion, ether injection, and ethanol injection. They are more cumbersome and are practically never used for the preparation of cationic liposomes. We shall mention only the detergent depletion methods which may be useful if some ligand-bearing lipids or proteins are to be included or embedded in the membrane. The lipid mixture is solubilized by a detergent with high aqueous solubility (critical micelle concentration). The components which are to be included are also dissolved in the detergent solution. Detergent is then removed by dialysis, gel chromatography, or by using detergent-adsorbing beads. This results in the formation of unilamellar vesicles. If octyl glucoside is used liposomes are above 100 nm while the use of sodium cholate results in the formation of SUVs. While this method doesn’t seem promising for large-scale liposome preparation, it can be used in conjunction with DNA to coat DNA–lipid particles with appropriate bilayers. Virosomes, vesicles containing viral lipids and some viral proteins, are typically produced by such a method also.
The Integrin α 6β4 in Epithelial and Carcinoma Cells
Published in Yoshikazu Takada, Integrins: The Biological Problems, 2017
Vito Quaranta, Ginetta Collo, Carla Rozzo, Lisa Starr, Guido Gaietta, Richard N. Tamura
In a diffèrent approach for identifying the α6β4 ligand, we prepared affinity columns with purified EHS laminin,57,58 a major component of basement membranes. We then passed over these columns surface-radioiodinated cell lysates from carcinoma cells, in order to determine whether α6β4 would bind. These lysates were prepared in the detergent octyl-glucoside, previously shown to be compatible with integrin binding to ligands.59 In several attempts, no α6β4 could be recovered from the laminin columns. Importantly, at least one β1 integrin, presumably one of the β1 laminin receptors,43 was eluted, suggesting that, under our conditions, the interaction of at least some integrins with their ligand is possible (R. N. Tamura, unpublished). From these experiments, we concluded that laminin (at least its EHS isoform57) is not the ligand for α6β4. This conclusion is in agreement with results from another laboratory,60 but is the exact opposite of the message from a recent report by Mercurio and co-workers,61 indicating that laminin (EHS isoform) is a ligand for α6β4. This conflict in the data is unresolved as yet, but could have several explanations. One important detail is that laminin binding of α6β4 was observed only with detergent extracts from one human cell line, Clone A, derived from a poorly differentiated adenocarcinoma of the colon, and said to “adhere avidly” to laminin.61 Besides trivial differences in affinity chromatography methods and ligand purification protocols, it could be that the state of activation of α6β4 in the particular cell line Clone A is such that laminin binding becomes detectable. State of activation may be the result of mutations in any of the subunits, or unknown intracellular or membrane factors, as seen for other integrins.62 The physiological significance of this result for other cell types remains to be proven.
Crystallizing Membrane Proteins: Experiments on Different Systems
Published in Hartmut Michel, Crystallization of Membrane Proteins, 1991
The design of our initial vapor diffusion experiments almost always resulted in, first, phase separation and then crystallization.5 Thus, we had concluded that the phase separation phenomenon was obligatory for crystallization. However, upon our development of a microdialysis system for porin crystallization, it became clear that crystals could be obtained without phase separation but only when the conditions were relatively close to the phase separation boundary.2 The microdialysis system allowed us to explore, in a systematic manner, the influence of each component on the system. We observed several distinct solute effects.2 First, moderate changes on the ionic strength were the primary determinant in selecting which of three crystal forms would grow. Second, changes in detergent concentration affected only the speed of nucleation and crystal growth, not crystal habit or quality. The slowest, most uniform crystal growth conditions occurred when the detergent concentration was only about 30% higher than the CMC. Third, the sensitivity of crystal growth and integrity to changes in temperature depended on the detergent used. For β-octyl glucoside, crystal nucleation and growth speed increased markedly with decreasing temperature. This effect can be reduced and even reversed by using noncarbohydrate based detergents (e.g., C8E5). The details of these effects strongly suggest that detergent-detergent interactions play a critical role in crystal nucleation and growth. This led us to use controlled temperature changes and detergent mixtures to fine-tune the crystallization process. Lastly, certain detergents and detergent mixtures could override the selection of the three primary crystal forms by solution ionic strength. In Figure 3c, triclinic crystals appeared when β-octyl glucoside was replaced with octyl 2-hydroxyethylsulfoxide. If detergents with dodecyl alkyl tails were used, the tetragonal to hexagonal transition could be induced by doping dode-cyldemethylamine oxide (LDAO) with C12E6. These experiments suggested to us that the nature of the detergent head group could profoundly affect how the PDC could pack in a crystal. Hence, the detergent surface on a PDC participates, in some manner, in the crystallization process.
Advances in lyotropic liquid crystal systems for skin drug delivery
Published in Expert Opinion on Drug Delivery, 2020
Ana Vitoria Pupo Silvestrini, Angelo Luis Caron, Juliana Viegas, Fabíola Garcia Praça, Maria Vitoria Lopes Badra Bentley
LLCs in situ have good cargo stability and system mechanical properties that make them promising candidates for drug administration in tissue regeneration. In this context, Wang et al. (2019) [89] demonstrated that the cubic LLC precursor containing VEGF promoted increased proliferation and migration in vitro. In vivo, angiogenesis was considerably increased 14 days after subcutaneous injection of the precursor LLC. The addition of octyl glucoside, a hydration-modulating agent, has been shown to increase the size of the cubic phase water channel, resulting in a sevenfold increase in VEGF release [89]. In tissue regeneration in chronic wounds, recombinant human epidermal growth factor (rhEGF)-containing LLC precursors incorporated with different proportions of ethanol, N-N-dimethylacetamide (DMAC) or PEG400 exhibited distinct viscosity, gelling, and release properties. In vivo studies with optimal formulations have shown promising effects on wound closure, immunomodulation, and re-epithelialization processes [214]. Another example of the application of LLC bulk in wound healing was proposed by Chen et al. (2020) [215]. The authors designed a spray dressing based on LLC combined with hyaluronic acid loaded with the anti-fibrotic drug pirfenidone. The application of the gel in situ provided excellent mechanical protection to promote healing and constant release of the asset, in addition to a control in the regulation of scar prophylaxis markers [215].
Factors affecting the morphology of some organic and inorganic nanostructures for drug delivery: characterization, modifications, and toxicological perspectives
Published in Expert Opinion on Drug Delivery, 2020
Ruchira Raychaudhuri, Abhjieet Pandey, Aswathi Hegde, Shaik Mohammad Abdul Fayaz, Dinesh Kumar Chellappan, Kamal Dua, Srinivas Mutalik
Silicone-based gemini surfactants of type Cm-PSi-Cm and hydrophobic chain of various lengths were synthesized and the properties of the respective surfactants were investigated [32]. A series of gemini surfactants with chain lengths of m = 8,10,12,14,16,18 were synthesized. The structure of the C12-PSi-C12 surfactant vesicles changed from micelles to vesicles and then progressed to rod-like, dumbbell-shaped, and finally to dumbbells arranged on a string, with the increase in the concentration of the surfactant. Morphological changes in mixtures of phosphatidylcholine-sodium cholate cause a transition from vesicle to micelles [33]. The initially spherical unilamellar vesicles lose their spherical nature upon cholate addition, which causes thread-like protrusions to emerge from the surface of the vesicle. When the concentration of cholate is increased, two phenomena happen parallelly. One is the detachment of the protrusions and the other is the destabilization of the bilayers. Ultimately, vesicles fragment into thread-like micelles. A system comprised C12PAM, a hydrophobically modified polyacrylamide and two oligomeric surfactants DTAD and PAHB showed a dependence of aggregate structure on surfactant concentration [34]. The changes in both the systems were similar from a soluble aggregate to a precipitate and again to the soluble phase. At very low surfactant concentrations, network-like aggregates were soluble. An increase in surfactant caused a formation of dense and cross-linked structures which form a precipitate. Without C12PAM, the surfactants alone could not aggregate and remain loosely arranged. Thus the interaction between C12PAM and DTAD/PAHB is the reason for the formation of aggregates. An additional increase in surfactant concentration further increases hydrophobic interactions and electrostatic repulsions, as a result, spherical aggregates are formed. The morphology of nanoparticle clusters embedded in bilayers of liposomes was found to depend on the surfactant concentration [35]. Oleic acid-coated SPIONs in aqueous buffer with β-octyl glucoside as a surfactant was mixed with liposomes and after removal of the surfactant, clusters of nanoparticles embedded in liposomes were obtained. At surfactant concentrations below its critical micelle concentration (CMC), liposomes formed were giant unilamellar vesicles and irregular aggregates were seen to be formed. When the concentration of β-octyl glucoside was adjusted close to its critical micelle concentration, showed no significant improvement as well. Above the critical micelle concentration, entire assembly of nanoparticle embedded liposomes took place from independent solubilized micelles.