Nanoscale Characterization of Nanomaterial - Cell Interactions
EMSL Project ID
25390
Abstract
This proposal is a continuation of research pursued under EMSL User Proposal #17690. The extensive use of amorphous silica nanoparticles (NPs) in the industry, and the growing use of carbon nanotubes (NTs), have caused the two manufactured nanomaterials to be a significant source for airborne environmental exposures. The two nanomaterials, which are fundamentally different in their shape and surface properties, impose adverse effects, but the underlying mechanism is far from being understood. The main target of exposure is the cells that line the respiratory tract, among them are the alveolar type II epithelial cells. These cells have apical microvilli that increase their surface area and vulnerability to the exposure. Whether and how amorphous silica NPs or carbon NTs penetrate these cells and what is the fate of the particle and the cell is unknown. Furthermore, the physical and chemical properties that govern these processes are unclear. Here we propose to investigate the cellular interactions, internalization pathways and fate of individual amorphous silica NPs and carbon NTs, while modifying their surface properties. Focusing on cultured alveolar type II epithelial cells, these studies will identify properties of inhaled nanomaterials that will increase their biocompatibility, and will aid in formulating preventative approaches. These studies will directly contribute to EMSL mission in supporting environmental research, and target EMSL Biological Interactions and Dynamics science theme by deciphering the involved protein interactions and cellular machinery dynamics. Accumulating observations suggest that nanomaterials exert harmful effects on human health to a greater extent than the larger particles, but the underlying mechanism is unclear. The harmful effect has been attributed, in part, to the large surface-area-to-mass ratio, leading to increased reactivity and oxidative stress. Under experimental conditions, where the localized concentration is high, nanomaterials tend to agglomerate, forming larger particles that can be detected and phagocytosed by the alveolar macrophages. However, it is currently thought that nanomaterials are able to escape the alveolar macrophages, and might directly enter the alveolar wall via the epithelial cells. Nanomaterials are therefore likely to be presented to cells in vivo as individual particles or small aggregates (<100 nm). The mechanisms of internalization and the cellular fate of individual or small nanomaterial aggregates are largely unknown. We will take advantage of EMSL unique capabilities in single-molecule fluorescence imaging techniques to identify individual or small clusters of nanomaterials and follow their spatial and temporal motion pattern in real time. Furthermore, EMSL capabilities in high resolution quantitative fluorescence resonance energy transfer (FRET) imaging and spectroscopy will be used to identify the underlying cellular interactions. Building on our recent findings that positively charged submicron and nanoscale amorphous silica particles are able to be propelled along the surface of filopodia and microvilli-like structures toward the cell body, we will identify the underlying protein interactions and cellular machinery. Using multi-channel fluorescence imaging, we will identify the endocytic pathways of individual nanomaterials, follow their trafficking within the cytoplasm and determine their final destination. Since the cellular interactions and pathways of nanomaterials are going to drive the fate of the cell and ultimately the level of toxicity, the new information will identify nanomaterial properties that can dictate biocompatibility.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2007-06-15
End Date
2010-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
6 Orr G, Panther DJ, Cassens KJ, Phillips JL, Tarasevich BJ, Pounds JG. Syndecan-1 mediates the coupling of positively charged submicrometer amorphous silica particles with actin filaments across the alveolar epithelial cell membrane. Toxicology and Applied Pharmacology. 2009, 236, 210-220.
6 Orr G, Panther DJ, Cassens KJ, Phillips JL, Tarasevich BJ, Pounds JG. Syndecan-1 mediates the coupling of positively charged submicrometer amorphous silica particles with actin filaments across the alveolar epithelial cell membrane. Toxicology and Applied Pharmacology. 2009, 236, 210?220.
G Orr, DJ Panther, JL Phillips, BJ Tarasevich, D Hu, JG Teeguarden, JG Pounds. (2007) Submicron and nanoscale inorganic particles exploit the actin machinery to be propelled along microvilli-like structures into alveolar cells. ACS Nano. 2007, 1(5)463-475.
Hess H and Tseng Y. “Active Intracellular Transport of Nanoparticles: Opportunity or Threat?” ACS Nano. 2007, 1(5), 390–392.
Orr G, WB Chrisler, KJ Cassens, R Tan, BJ Tarasevich, LM Markillie, RC Zangar, and BD Thrall. 2010. "Cellular Recognition and Trafficking of Amorphous Silica Naoparticles by Macrophage Scavenger Receptor A." Nanotoxicology. [In Press]
Teeguarden JG, PM Hinderliter, G Orr, BD Thrall, and JG Pounds. (2007) Particokinetics In Vitro: Dosimetry Considerations for In Vitro Nanoparticle Toxicity Assessments. Toxicological Science. Feb;95(2):300-12. Epub 2006 Nov 10.
Waters KM, LM Masiello, RC Zangar, BJ Tarasevich, NJ Karin, RD Quesenberry, S Bandyopadhyay, JG Teeguarden, JG Pounds, and BD Thrall. 2009. "Macrophage Responses to Silica Nanoparticles are Highly Conserved Across Particle Sizes." Toxicological Sciences 107(2):553-569.