Examining the Cellular Uptake and Toxicity of Engineered Nanomaterials

Wednesday, May 14, 2014: 09:00
Bonnet Creek Ballroom VI, Lobby Level (Hilton Orlando Bonnet Creek)
K. Kosaraju (University of North Carolina at Greensboro), M. Tarannum, S. Crawford, K. Garde, and S. Aravamudhan (North Carolina A&T State University)
Nanomaterials have attracted great attention for numerous applications in chemical, biological, and industrial world because of their fascinating physicochemical properties. In particular, understanding the mechanism of engineered nanomaterials (ENs) uptake by cells is important for various biomedical applications including for biosensors, imaging, intracellular drug and gene delivery, and toxicity studies (Han et al. 2007). Spontaneous penetration of functionalized cationic Au NPs has shown cell membrane disruption and cytotoxicity, thus limiting their utility (Chen et al. 2009). The objective of this work is to study and quantify the cellular uptake of three types of ENs (CNTs, Au nanoparticles (NPs), Silica and ZnO NPs) and their corresponding mechanism of action using immunofluorescence staining, ultrastructural characterization (HRTEM), Electrochemical Cell-substrate Impedance Sensing (ECIS) and bioanalytical analysis (ICP-OES). Another recent literature have indicated that Au NPs (5 nm diameter with “special” surface chemistries or arrangements) protected by an amphiphilic monolayer can non-disruptively penetrate cell membrane to deliver drugs, nutrients or biosensors (Verma et al. 2008). Although it seems that ENs can be taken up by cells, the evidence is disparate and the mechanism of uptake is either unclear or in their infancy. This is because of lack of accurate data on the physicochemical properties such as size including surface area, size distribution, chemical composition (purity crystallinity, electronic properties, etc.), solubility, shape and aggregation; and surface structure including surface reactivity, groups and protein corona and their relationship to cellular uptake. Of particular importance is the protein corona, which affects how nanoparticles are internalized by cells and cleared from the body. When NPs are exposed to biological fluid such as media in a cell culture, proteins can bind to the surface of the nanoparticle to form a protein corona. In this study, we first comprehensively characterized the starting EN dispersed in cell media (using DLS, BET, FTIR, Raman, XRD and HRSEM). Next, NPs were exposed to different cell types (A549, NIH 3T3 and PC12 cells) for either 1 or 2 days.  The goal here is to address three questions – (a) Do ENs penetrate into cells and how does intracellular trafficking of ENs occur? (b) Does uptake of ENs vary with cell type? (c) Is uptake driven by physiochemical factors such as size, aggregation, shape, functionalization, corona, composition, dose and time? Figure 1 shows typical studies of cellular uptake of EN analyzed using VP-SEM, ICP-OES, immunofluorescence, and ECIS. Our initial evidence shows nanoparticle transport and uptake (see ICP-OES) and cell morphological disruptions. Furthermore, ECIS studies indicate the ability to longitudinally monitor impedance data of exposed cells over 5 days. In the paper, we will present detailed investigation on the influence of other EN physicochemical properties on cellular transport and uptake.