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Chen, Ran
Cell; Membrane; Nanoparticle; Physicochemistry; Protein corona; Uptake/discharge; Physics
thesis / dissertation description
Nanotechnology has revolutionalized the landscape of modern science and technology, including materials, electronics, therapeutics, bioimaging, sensing, and the environment. Along with these technological advancements, there arises a concern that engineered nanomaterials, owing to their high surface area and high reactivity, may exert adverse effects upon discharge to compromise biological and ecological systems. Research in the past decade has examined the fate of nanomaterials in vitro and in vivo, as well as the interactions between nanoparticles and biological and ecosystems using primarily toxicological and ecotoxicological approaches. However, due to the versatility in the physical and physicochemical properties of nanoparticles, and due to the vast complexity of their hosting systems, the solubility, transformation, and biocompatibility of nanomaterials are still poorly understood. Accordingly, this dissertation offers a mechanistic study on the differential translocation of pristine and water-soluble fullerene nanoparticles in mammalian and plant cells (Chapter 2), an investigation on membrane fluidity upon exocytosis of gold nanoparticles by the cell (Chapter 3), and an in-depth examination of the formation of an array of nanoparticle-protein coronas and their interactions with lipid vesicles and the cell (Chapters 4 and 5). The organization of this dissertation is as follows. Chapter 1 presents a review on the general applications (gene and drug delivery, imaging, sensing, nanotherapy) and implications (toxicity) of nanomaterials, mostly within the context of biological systems. In addition, this chapter documents theendocytotic and exocytotic pathways of the cell, and reviews the state-of-the-art of our understanding of nanoparticle-protein corona formation and nanoparticle-cell interactions, two precursors of nanotoxicity. Chapter 2 offers, for the first time, a parallel study on the differential uptake of hydrophobic and amphiphilic fullerene nanoparticles by Allium cepa plant cells and HT- 29 mammalian cells, two model systems representing ecological and biological systems. Methodologically, this study was conducted using a plant cell viability assay, bright field and fluorescence imaging, and, extensively, electron microscopy imaging. Chapter 3 examines an important but rarely documented aspect of cellular response to nanoparticles - exocytosis. A lipophilic Laurdan dye was used to partition into HT-29 mammalian cell membranes. Membrane fluidity as a result of the discharge of gold nanoparticles was inferred from UV-vis absorbance as well as by calculating the general polarization value of the dye -- hereby treated an electric dipole in a lipid bilayer continuum -- based on its fluorescence emissions at two characteristic wavelengths. Chapter 4 concerns protein adsorption on carbon nanotubes (CNTs) to form protein coronas in cell culture media, an environment relevant to both in vitro and in vivo studies. A label-free mass spectrometry-based proteomic approach was employed, and the compositions of the protein forming coronas on a set of CNTs were examined. The physicochemical properties of the CNTs were also extensively characterized in order to establish a correlation between protein adsorption and CNT surface properties. Chapter 5 characterizes the formation of a serum albumin corona on silver nanoparticles and evaluates the impact of silver nanoparticle-albumin corona on thefluidity of an artificial lipid vesicle. The reason of adopting a lipid vesicle in this study is to eliminate endo- and exocytosis and pinpoint the roles of physical forces in nanoparticle-cell interactions. In this chapter we also show the formation and conformational changes of fibrinogen corona in HT-29 cell lines. Fibrinogen is one of the most abundant types of plasma proteins in the bloodstream. Chapter 6 summarizes the major findings in this dissertation and presents future work inspired by this Doctor of Philosophy (PhD) research.