The influence of shape and charge on protein corona composition in common gold nanostructures.

With increasing importance of gold nanoparticles (AuNPs) in the medical field, the understanding of their interactions in biological environments is essential. It is known that the exposure to biological fluids of particles in the nanometric range leads to accumulation of proteins on the particle surface proximity, generating the so-called protein corona. This fact can completely change the properties of AuNPs, thus drastically influencing the characteristics and intended purpose of the particles. Therefore, deep insight on the formation and composition of this protein corona is of extreme importance. Between the different factors that can alter the corona formation, our study focuses on the influence of the shape and particle surface charge. In detail, four different shapes of nanometrical scale (spheres, rods, stars and cages) of comparable size were used, all of them stabilized with three different heterofunctionalized poly(ethylene glycol) thiol (R-PEG-SH) linkers (R = OCH3, COOH or NH2) to check the effect of charge as well. After incubation with human serum, abundant proteins were identified via liquid chromatography-electrospray ionization-tandem mass spectroscopy (LC ESI MS/MS) and compared in terms of their relative abundance. On the basis of statistical evaluations, the shape of our AuNPs showed a greater influence than the surface charge. Especially, cage-shaped AuNPs showed a lower amount of total corona proteins. This shape showed differences in the abundances of individual proteins like albumin, vitronectin and members of the complement system. These results indicate that nanocages could present an improved biocompatibility compared with the other shapes due to the high curvature areas and dense ligation on the flat surfaces that could hinder opsonisation and fast removal by the immune system.

Amphiphilic nanogels: influence of surface hydrophobicity on protein corona, biocompatibility and cellular uptake.

BACKGROUND AND PURPOSE: Nanogels (NGs) are promising drug delivery tools but are typically limited to hydrophilic drugs. Many potential new drugs are hydrophobic. Our study systematically investigates amphiphilic NGs with varying hydrophobicity, but similar colloidal features to ensure comparability. The amphiphilic NGs used in this experiment consist of a hydrophilic polymer network with randomly distributed hydrophobic groups. For the synthesis we used a new synthetic platform approach. Their amphiphilic character allows the encapsulation of hydrophobic drugs. Importantly, the hydrophilic/hydrophobic balance determines drug loading and biological interactions. In particular, protein adsorption to NG surfaces is dependent on hydrophobicity and critically determines circulation time. Our study investigates how network hydrophobicity influences protein binding, biocompatibility and cellular uptake. METHODS: Biocompatibility of the NGs was examined by WST-1 assay in monocytic-like THP-1 cells. Serum protein corona formation was investigated using dynamic light scattering and two-dimensional gel electrophoresis. Proteins were identified by liquid chromatography-tandem mass spectrometry. In addition, cellular uptake was analyzed via flow cytometry. RESULTS: All NGs were highly biocompatible. The protein binding patterns for the two most hydrophobic NGs were very similar to each other but clearly different from the hydrophilic ones. Overall, protein binding was increased with increasing hydrophobicity, resulting in increased cellular uptake. CONCLUSION: Our study supports the establishment of structure-property relationships and contributes to the accurate balance between maximum loading capacity with low protein binding, optimal biological half-life and good biocompatibility. This is an important step to derive design principles of amphiphilic NGs to be applied as drug delivery vehicles.

The influence of surface charge on serum protein interaction and cellular uptake: studies with dendritic polyglycerols and dendritic polyglycerol-coated gold nanoparticles.

Nanoparticles (NPs) have gained huge interest in the medical field, in particular for drug delivery purposes. However, binding of proteins often leads to fast NP uptake and rapid clearance, thereby hampering medical applications. Thus, it is essential to determine and control the bio-nano interface. This study investigated the serum protein interactions of dendritic polyglycerols (dPGs), which are promising drug delivery candidates by means of two dimensional gel electrophoresis (2DE) in combination with mass spectrometry. In order to investigate the influence of surface charge, sulfated (sulfated dendritic polyglycerol [dPGS]) and non-sulfated (dPGOH) surfaces were applied, which were synthesized on a gold core allowing for easier separation from unbound biomolecules through centrifugation. Furthermore, two different sizes for dPGS were included. Although size had only a minor influence, considerable differences were detected in protein affinity for dPGS versus dPGOH surfaces, with dPGOH binding much less proteins. Cellular uptake into human CD14+ monocytes was analyzed by flow cytometry, and dPGOH was taken up to a much lower extent compared to dPGS. By using a pull-down approach, possible cellular interaction partners of serum pre-incubated dPGS-Au20 NPs from the membrane fraction of THP-1 cells could be identified such as for instance the transferrin receptor or an integrin. Clathrin-mediated endocytosis was further investigated using chlorpromazine as an inhibitor, which resulted in a 50% decrease of the cellular uptake of dPGS. This study could confirm the influence of surface charge on protein interactions and cellular uptake of dPGS. Furthermore, the approach allowed for the identification of possible uptake receptors and insights into the uptake mechanism.