Internalization of SiO2 nanoparticles by alveolar macrophages and lung epithelial cells and its modulation by the lung surfactant substitute Curosurf®

Nanomaterials (NM) exhibit novel physicochemical properties that determine their interaction with biological substrates and processes. Three metal oxide nanoparticles that are currently being produced in high tonnage, TiO(2), ZnO, and CeO(2), were synthesized by flame spray pyrolysis process and compared in a mechanistic study to elucidate the physicochemical characteristics that determine cellular uptake, subcellular localization, and toxic effects based on a test paradigm that was originally developed for oxidative stress and cytotoxicity in RAW 264.7 and BEAS-2B cell lines. ZnO induced toxicity in both cells, leading to the generation of reactive oxygen species (ROS), oxidant injury, excitation of inflammation, and cell death. Using ICP-MS and fluorescent-labeled ZnO, it is found that ZnO dissolution could happen in culture medium and endosomes. Nondissolved ZnO nanoparticles enter caveolae in BEAS-2B but enter lysosomes in RAW 264.7 cells in which smaller particle remnants dissolve. In contrast, fluorescent-labeled CeO(2) nanoparticles were taken up intact into caveolin-1 and LAMP-1 positive endosomal compartments, respectively, in BEAS-2B and RAW 264.7 cells, without inflammation or cytotoxicity. Instead, CeO(2) suppressed ROS production and induced cellular resistance to an exogenous source of oxidative stress. Fluorescent-labeled TiO(2) was processed by the same uptake pathways as CeO(2) but did not elicit any adverse or protective effects. These results demonstrate that metal oxide nanoparticles induce a range of biological responses that vary from cytotoxic to cytoprotective and can only be properly understood by using a tiered test strategy such as we developed for oxidative stress and adapted to study other aspects of nanoparticle toxicity.

In this work, we explore the formation of the protein corona after exposure of metallic Au nanoparticles (NPs), with sizes ranging from 4 to 40 nm, to cell culture media containing 10% of fetal bovine serum. Under in vitro cell culture conditions, zeta potential measurements, UV-vis spectroscopy, dynamic light scattering and transmission electron microscope analysis were used to monitor the time evolution of the inorganic NP-protein corona formation and to characterize the stability of the NPs and their surface state at every stage of the experiment. As expected, the red-shift of the surface plasmon resonance peak, as well as the drop of surface charge and the increase of the hydrodynamic diameter indicated the conjugation of proteins to NPs. Remarkably, an evolution from a loosely attached toward an irreversible attached protein corona over time was observed. Mass spectrometry of the digested protein corona revealed albumin as the most abundant component which suggests an improved biocompatibility.

The importance of the protein corona formed around nanoparticles upon entering a biological fluid has recently been highlighted. This corona is, when sufficiently long-lived, thought to govern the particles' biological fate. However, even this long-lived "hard" corona evolves and re-equilibrates as particles pass from one biological fluid to another, and may be an important feature for long-term fate. Here we show the evolution of the protein corona as a result of transfer of nanoparticles from one biological fluid (plasma) into another (cytosolic fluid), a simple illustrative model for the uptake of nanoparticles into cells. While no direct comparison can be made to what would happen in, for example, the uptake pathway, the results confirm that significant evolution of the corona occurs in the second biological solution, but that the final corona contains a "fingerprint" of its history. This could be evolved to map the transport pathways utilized by nanoparticles, and eventually to predict nanoparticle fate and behavior. © 2011 American Chemical Society