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• W2592037027 abstract "This thesis presents different approaches for the surface engineering by means of polyelectrolyte multilayers (PEMs), alone or in combination with lipid bilayers and influenza virosomes for biomedical applications.In chapter 1, PEMs of poly-L-lysine (PLL) and alginic acid sodium salt (Alg) are fabricated applying the layer by layer (LbL) technique and annealed at constant temperatures; 37, 50 and 80 °C, for 72 hours. Atomic force microscopy (AFM) reveals changes in the topography of the PEM, which is changing from a fibrillar to a smooth surface after annealing. Advancing contact angle in water varies from 36° before annealing to around 93°, 77° and 95° after annealing at 37, 50 and 80 °C respectively. Changes in surface energy after annealing were calculated from advancing and receding contact angle measurements performed with water and with organic solvents. Changes in the physical properties of the PEMs are interpreted as a result of the reorganization of the polyelectrolytes in the PEMs from a layered structure into complexes where the interaction of polycations and polyanions is enhanced. PEMs from biological origin have many potential applications in tissue engineering and regenerative medicine. However, the softness of biocompatible PEMs results in limited cell adhesion. Thermal annealing is suggested as a novel strategy for the enhancement of cellularadhesion on PEMs. The impact of thermal annealing at 37 °C, on the adhesion of human lung cancer A549 and myoblast C2C12 cell lines is studied. Cell adhesion, measured by the projected average cells spreading and focal contact is remarkably improved for the annealed PEMs. Quartz crystal microbalance with dissipation (QCM-D), contact angle and fluorescence spectroscopy measurements show a significant decrease in the adsorption of the bovine serum albumin protein to the PEMs after annealing. Our results provide a simple method to tune the wettability of bio-PEMs, improve cellular adhesion and endow them with antifouling characteristics.In chapter 2, the self-assembly of small unilamellar vesicles (SUVs) of mixed lipids zwitterionic phosphatidylcholine (DOPC, PC) and anionic phosphatidylserine (DOPS, PS) phospholipids on top of PEMs of poly(allylamine hydrochloride) (PAH), as a polycation, and poly(sodium 4-styrenesulfonate) (PSS), as a polyanion, is investigated as a function of the composition of the assembled vesicles by means of QCM-D, cryo-transmission electron microscopy (CryoTEM), AFM and atomic force spectroscopy (AFS). Vesicles with molar percentages of PS between 50 % and 70 % result in the formation of a lipid bilayer on top of the PEMs. AFS studies performed with a PAH-modified cantilever approaching and retracting from the lipid assemblies reveal that the main interaction between PAH and the lipids takes place through hydrogen bonding between the amine groups of PAH and the carboxylate and phosphate groups of PS and with the phosphate groups of PC.The influence of the surface chemistry of PEMs on the formation of lipid bilayers is also studied for PEMs with poly(diallyldimethylammonium chloride) (PDADMAC) aspolycation and top layer, and PSS as polyanion. SUVs composed of DOPC and DOPS at 50:50 molar ratio are deposited on top of PEM films and studied via QCM-D and fluorescence recovery after photobleaching (FRAP). SUVs deposition on PDADMAC/PSS results in vesicles adsorption while on PAH/PSS under the same conditions a bilayer is formed. FRAP measurements confirm that SUVs are not ruptured on top of PDADMAC/PSS. The role of phosphate ions, in solution, on the formation of lipid bilayers is also analysed. ?-ray photoelectron spectroscopy (XPS) shows the complexation of phosphate salts to the primary amines of PAH and no interaction with the quaternary amines of PDADMAC. ? ? potential measurements show a potential close to 0 mV for the PAH/PSS multilayers in PBS while PDADMAC/PSS display a potential of 38 mV. A model is presented for the formation of lipid bilayers on PAH/PSS PEMs taking into account the role of phosphate ions in decreasing electrostatic interactions between SUVs and PEMs and the formation of hydrogen bonding between the phospholipids and the primary amines of PAH.QCM-D and FRAP experiments show that when vesicles with a lipid composition of 50:50 DOPC:DOPS are adsorbed on PEMs where PSS is replaced by Alg or poly(acrylic acid) (PAA) the vesicle deposition does not result in a bilayer formation but in bilayer patches together with adsorbed intact vesicles. Therefore, the fusion of the lipid bilayer is not only affected by the last deposited layer that mainly interacts with the lipids but also by the overall composition of the PEM film.In chapter 3, SUVs prepared by a mixture of 30:70 DOPC:DOPS are assembled on top of a PEM cushion of PAH/PSS and the electrical properties of the bilayer are studied byelectrochemical impedance spectroscopy (EIS). The bilayer supported on the PEMs shows a high resistance, in the order of 107 ? cm2 which is indicative of a continuous, dense bilayer. Such resistance is comparable with the resistance of black lipid membranes, being the first time that these values are obtained for lipid bilayers supported on PEMs. The assembly of polyelectrolytes on top of a lipid bilayer decreases the resistance of the bilayer up to 2 orders of magnitude. Thus the assembly of the polyelectrolytes on the lipid bilayer induces defects or pores in the bilayer followed by a subsequent decrease in resistance.Finally, this thesis addresses, in Chapter 4, the fusion of immunostimulating reconstituted influenza virosomes (IRIVs) with the functional viral envelope glycoprotein, hemagglutinin (HA), to artificial supported lipid membranes assembled on PAH/PSS PEMs on both colloidal particles and planar substrates. R18 assay is used to prove the IRIVs fusion in dependence of pH, temperature and HA concentration. IRIVs display a pH-dependent fusion mechanism, fusing at low pH in analogy to the influenza virus. The pH dependent behaviour is confirmed by QCM-D. AFM imaging shows that at low pH virosomes are integrated in the supported membrane displaying flatered features and a reduced vertical thickness. IRIVs fusion offers a new strategy for transferring biological functions on artificial supported membranes with potential applications in targeted delivery and sensing." @default.
• W2592037027 created "2017-03-16" @default.
• W2592037027 creator A5005160351 @default.
• W2592037027 date "2016-12-19" @default.
• W2592037027 modified "2023-09-23" @default.
• W2592037027 title "Engineered interfaces with polyelectrolyte multilayers, lipid bilayer membranes and virosomes for biomedical applications" @default.
• W2592037027 hasPublicationYear "2016" @default.
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