Patrick N. Sisco

Transfer of gold nanoparticles from the water column to the estuarine food web.

Within the next five years the manufacture of large quantities of nanomaterials may lead to unintended contamination of terrestrial and aquatic ecosystems. The unique physical, chemical and electronic properties of nanomaterials allow new modes of interaction with environmental systems that can have unexpected impacts. Here, we show that gold nanorods can readily pass from the water column to the marine food web in three laboratory-constructed estuarine mesocosms containing sea water, sediment, sea grass, microbes, biofilms, snails, clams, shrimp and fish. A single dose of gold nanorods (65 nm length x 15 nm diameter) was added to each mesocosm and their distribution in the aqueous and sediment phases monitored over 12 days. Nanorods partitioned between biofilms, sediments, plants, animals and sea water with a recovery of 84.4%. Clams and biofilms accumulated the most nanoparticles on a per mass basis, suggesting that gold nanorods can readily pass from the water column to the marine food web.

Surface-Coverage Dependence of Surface-Enhanced Raman Scattering from Gold Nanocubes on Self-Assembled Monolayers of Analyte†

Surface enhanced Raman scattering (SERS) spectra of 4-mercaptobenzoic acid (4-MBA) self-assembled monolayers (SAMs) on gold substrates are presented for SAMs onto which gold nanocubes have been electrostaticaly immobilized. In the absence of nanocubes, no SERS signals from 4-MBA SAMs are observed. Upon addition of the gold nanocubes to the SAM, a sandwich architecture is formed, allowing for coupling between the localized surface plasmon of the nanocubes and the surface plasmon of the gold substrate. This creates a large electromagnetic field in the area where the 4-MBA molecules reside, causing the characteristic vibrational modes of 4-MBA to appear. SERS intensities increase linearly with increasing nanocube coverage up to a factor of 7 in the best case studied here, with enhancement factors of up to 10(13).

Polyelectrolyte-coated gold nanorods and their interactions with type I collagen

Gold nanorods (AuNRs) have unique optical properties for numerous biomedical applications, but the interactions between AuNRs and proteins, particularly those of the extracellular matrix (ECM), are poorly understood. Here the effects of AuNRs on the self-assembly, mechanics, and remodeling of type I collagen gels were examined in vitro. AuNRs were modified with polyelectrolyte multilayers (PEMs) to minimize cytotoxicity, and AuNRs with different terminal polymer chemistries were examined for their interactions with collagen by turbidity assays, rheological tests, and microscopy. Gel contraction assays were used to examine the effects of the PEM-coated AuNRs on cell-mediated collagen remodeling. Polyanion-terminated AuNRs significantly reduced the lag (nucleation) phase of collagen self-assembly and significantly increased the dynamic shear modulus of the polymerized gels, whereas polycation-terminated AuNRs had no effect on the mechanical properties of the collagen. Both polyanion- and polycation-terminated AuNRs significantly inhibited collagen gel contraction by cardiac fibroblasts, and the nanoparticles were localized in intra-, peri-, and extracellular compartments, suggesting that PEM-coated AuNRs influence cell behavior via multiple mechanisms. These results demonstrate the significance of nanoparticle-ECM interactions in determining the bioactivity of nanoparticles.

The effect of gold nanorods on cell-mediated collagen remodeling

Cardiac fibroblasts, the noncontractile cells of the heart, contribute to myocardial maintenance through the deposition, degradation, and organization of collagen. Adding polyelectrolyte-coated gold nanorods to three-dimensional constructs composed of collagen and cardiac fibroblasts reduced contraction and altered the expression of mRNAs encoding beta-actin, alpha-smooth muscle actin, and collagen type I. These data show that nanomaterials can modulate cell-mediated matrix remodeling and suggest that the targeted delivery of nanomaterials can be applied for antifibrotic therapies.

Adsorption of cellular proteins to polyelectrolyte-functionalized gold nanorods: a mechanism for nanoparticle regulation of cell phenotype?

Cell behavior in the presence of nanomaterials is typically explored through simple viability assays, but there is mounting evidence that nanomaterials can have more subtle effects on a variety of cellular functions. Previously our lab demonstrated that gold nanorods functionalized with polyelectrolyte multi-layers inhibited rat cardiac fibroblast-mediated remodeling of type I collagen scaffolds by altering fibroblast phenotype and the mechanical properties of the collagen network. In this work, we examine a possible mechanism for these effects: adsorption of cellular proteins by the nanorods. Mass spectrometric and gel electrophoresis of media collected from cultured cells suggests that a number of proteins, some of which mediate cell-cell and cell-matrix interactions, adsorb onto the surface of these nanoparticles in vitro. Polyethylene glycol coating of the nanorods largely mitigates protein adsorption and fibroblast-mediated collagen remodeling. These results suggest that adsorption of proteins by nanorods could have a significant effect on cell functions, including fibroblast-mediated matrix remodeling.

Glycosaminoglycan-functionalized gold nanorods: interactions with cardiac cells and type I collagen

The sulfated glycosaminoglycans (sGAG) heparin and chondroitin sulfate (CS) were immobilized on the surfaces of gold nanorods as part of a polyelectrolyte multilayer. The effects of these nanomaterials on the self-assembly of type I collagen were examined by turbidity assays and microscopy, and desorption of sGAG from nanomaterial-collagen composites was quantified biochemically. The interactions of sGAG-coated nanorods with cardiac cells were also explored through a collagen gel contraction assay and confocal microscopy. In contrast to soluble forms of sGAG, sGAG-coated nanorods consistently accelerated collagen fibrillogenesis. Soluble heparin, and heparin- and CS-coated nanorods inhibited cell-mediated contraction of collagen gels, whereas soluble CS did not. Both heparin and CS-coated nanorods were detected in the peri- and/or intra-cellular compartments of the cells, but there was no evidence of cytotoxicity over 72 h of culture. These results indicate that biological polyanions, such as sGAG, may be useful in the modification of nanoparticle surface chemistry for biological and/or therapeutic applications.

Surface Charge Controls the Fate of Au Nanorods in Saline Estuaries

This work reports the distribution of negatively charged, gold core nanoparticles in a model marine estuary as a function of time. A single dose of purified polystyrene sulfonate (PSS)-coated gold nanorods was added to a series of three replicate estuarine mesocosms to emulate an abrupt nanoparticle release event to a tidal creek of a Spartina -dominated estuary. The mesocosms contained several phases that were monitored: seawater, natural sediments, mature cordgrass, juvenile northern quahog clam, mud snails, and grass shrimp. Aqueous nanorod concentrations rose rapidly upon initial dosing and then fell to stable levels over the course of approximately 50 h, after which they remained stable for the remainder of the experiment (41 days total). The concentration of nanorods rose in all other phases during the initial phase of the experiment; however, some organisms demonstrated depuration over extended periods of time (100+ h) before removal from the dosed tanks. Clams and biofilm samples were also removed from the contaminated tanks post-exposure to monitor their depuration in pristine seawater. The highest net uptake of gold (mass normalized) occurred in the biofilm phase during the first 24 h, after which it was stable (to the 95% level of confidence) throughout the remainder of the exposure experiment. The results are compared against a previous study of positively charged nanoparticles of the same size to parameterize the role of surface charge in determining nanoparticle fate in complex aquatic environments.

High-aspect-ratio gold nanorods: their synthesis and application to image cell-induced strain fields in collagen films.

Gold nanoparticles are receiving considerable attention due to their novel properties and the potential variety of their uses. Long gold nanorods with dimensions of approximately 20 × 400 nm exhibit strong light scattering and can be easily observed under dark-field microscopy. Here we describe the use of this light-scattering property to track micrometer scale strains in collagen gels and thick films, which result from cell traction forces applied by neonatal heart fibroblasts. The use of such collagen constructs to model cell behavior in the extracellular matrix is common, and describing local mechanical environments on such a small scale is necessary to understand the complex factors associated with the remodeling of the collagen network. Unlike other particles used for tracking purposes, gold nanorods do not photobleach, allowing their optical signal to be tracked for longer periods of time, and they can be easily synthesized and coated with various charged or neutral shells, potentially reducing the effect of their presence on the cell system or allowing selective placement. Techniques described here are ultimately applicable for investigations with a wide variety of cells and cell environments.