Effect of the Protein Corona Formation on Antibody Functionalized Liquid Lipid Nanocarriers.

The main aim of this study is to report basic knowledge on how a protein corona (PC) could affect or modify the way in which multifunctionalized nanoparticles interact with cells. With this purpose, we have firstly optimized the development of a target-specific nanocarrier by coupling a specific fluorescent antibody on the surface of functionalized lipid liquid nanocapsules (LLNCs). Thus, an anti-HER2-FITC antibody (αHER2) has been used, HER2 being a surface receptor that is overexpressed in several tumor cells. Subsequently, the in vitro formation of a PC has been developed using fetal bovine serum supplemented with human fibrinogen. Dynamic Light Scattering (DLS), Nanoparticle Tracking Analysis (NTA), Laser Doppler Electrophoresis (LDE), and Gel Chromatography techniques have been used to assure a complete physico-chemical characterization of the nano-complexes with (LLNCs-αHER2-PC) and without (LLNCs-αHER2) the surrounding PC. In addition, cellular assays were performed to study the cellular uptake and the specific cellular-nanocarrier interactions using the SKBR3 (high expression of HER2) breast cancer cell line and human dermal fibroblasts (HDFa) (healthy cell line without expression of HER2 receptors as control), showing that the SKBR3 cell line had a higher transport rate (50-fold) than HDFa at 60 min with LLNCs-αHER2. Moreover, the SKBR3 cell line incubated with LLNCs-αHER2-PC suffered a significant reduction (40%) in the uptake. These results suggest that the formation of a PC onto LLNCs does not prevent specific cell targeting, although it does have an important influence on cell uptake.

Comparative interfacial in vitro digestion of protein and polysaccharide oil/water films.

The behaviour of proteins (ß-lactoglobulin (ßlg) and soy protein isolate (SPI)) and a surface active polysaccharide (hydroxypropylmethylcellulose, HPMC) o/w interfacial films under simulated gastrointestinal conditions using the interfacial tensiometer Octopus were compared and related to the performance of the emulsions (using the same emulsifiers) under in vitro digestion. The evolution of interfacial tension (γ) was used to investigate the effect of gastrointestinal fluids on o/w interfacial films. Clear differences were observed among these emulsifiers. During the gastric phase, HPMC showed the lowest change in γ values as compared to protein films. The most important changes occurred during the intestinal stage where it was observed an important decrease of γ associated with the rapid penetration of BS, followed by a lower rate of decrease attributable to the accumulation of FFA at the interface. In the last stage, the subphase was exchanged by buffer alone, to remove the reversibly adsorbed digestion products. SPI formed the most resistant interface to the remotion of digestion products, followed by HPMC and finally by ßlg. The results agree with the degree of lipolysis reported for the emulsions stabilized by these emulsifiers, which suggest that lipid digestion could be modulated by the ability of emulsifiers to prevent the BS activity (to adsorb at the O/W interface or remove the inhibitory digestion products from the interface). Thus, emulsifiers-BS interactions appears as a key factor in controlling the lipolysis.

Dual delivery nanosystem for biomolecules. Formulation, characterization, and in vitro release.

Because of the biocompatible and biodegradable properties of poly (lactic-co-glycolic acid) (PLGA), nanoparticles (NPs) based on this polymer have been widely studied for drug/biomolecule delivery and long-term sustained-release. In this work, two different formulation methods for lysozyme-loaded PLGA NPs have been developed and optimized based on the double-emulsion (water/oil/water, W/O/W) solvent evaporation technique. They differ mainly in the phase in which the surfactant (Pluronic® F68) is added: water (W-F68) and oil (O-F68). The colloidal properties of these systems (morphology by SEM and STEM, hydrodynamic size by DLS and NTA, electrophoretic mobility, temporal stability in different media, protein encapsulation, release, and bioactivity) have been analyzed. The interaction surfactant-protein depending on the formulation procedure has been characterized by surface tension and dilatational rheology. Finally, cellular uptake by human mesenchymal stromal cells and cytotoxicity for both systems have been analyzed. Spherical hard NPs are made by the two methods However, in one case, they are monodisperse with diameters of around 120nm (O-F68), and in the other case, a polydisperse system of NPs with diameters between 100 and 500nm is found (W-F68). Protein encapsulation efficiency, release and bioactivity are maintained better by the W-F68 formulation method. This multimodal system is found to be a promising "dual delivery" system for encapsulating hydrophilic proteins with strong biological activity at the cell-surface and cytoplasmic levels.