The rapid growth in the use of in vitro methods for nanoparticle toxicity assessment has proceeded with limited consideration of the unique kinetics of these materials in solution. Particles in general and nanoparticles specifically, diffuse, settle, and agglomerate in cell culture media as a function of systemic and particle properties: media density and viscosity and particle size, shape, charge and density, for example. Cellular dose then is also a function of these factors as they determine the rate of transport of nanoparticles to cells in culture. Here we develop and apply the principles of dosimetry in vitro and outline an approach for simulation of nanoparticle particokinetics in cell culture systems. We illustrate that where equal mass concentrations (mug/ml) imply equal doses for dissimilar materials, the corresponding particle number or surface area concentration doses differ by orders of magnitude. More importantly, when rates of diffusional and gravitational particle delivery are accounted for, trends and magnitude of the cellular dose as a function of particle size and density differ significantly from those implied by "concentration" doses. For example, 15-nm silver nanoparticles appear approximately 4000 times more potent than micron-sized cadmium oxide particles on a cm(2)/ml media basis, but are only approximately 50 times more potent when differences in delivery to adherent cells are considered. We conclude that simple surrogates of dose can cause significant misinterpretation of response and uptake data for nanoparticles in vitro. Incorporating particokinetics and principles of dosimetry would significantly improve the basis for nanoparticle toxicity assessment, increasing the predictive power and scalability of such assays.

BackgroundThe difficulty of directly measuring cellular dose is a significant obstacle to application of target tissue dosimetry for nanoparticle and microparticle toxicity assessment, particularly for in vitro systems. As a consequence, the target tissue paradigm for dosimetry and hazard assessment of nanoparticles has largely been ignored in favor of using metrics of exposure (e.g. μg particle/mL culture medium, particle surface area/mL, particle number/mL). We have developed a computational model of solution particokinetics (sedimentation, diffusion) and dosimetry for non-interacting spherical particles and their agglomerates in monolayer cell culture systems. Particle transport to cells is calculated by simultaneous solution of Stokes Law (sedimentation) and the Stokes-Einstein equation (diffusion).ResultsThe In vitro Sedimentation, Diffusion and Dosimetry model (ISDD) was tested against measured transport rates or cellular doses for multiple sizes of polystyrene spheres (20-1100 nm), 35 nm amorphous silica, and large agglomerates of 30 nm iron oxide particles. Overall, without adjusting any parameters, model predicted cellular doses were in close agreement with the experimental data, differing from as little as 5% to as much as three-fold, but in most cases approximately two-fold, within the limits of the accuracy of the measurement systems. Applying the model, we generalize the effects of particle size, particle density, agglomeration state and agglomerate characteristics on target cell dosimetry in vitro.ConclusionsOur results confirm our hypothesis that for liquid-based in vitro systems, the dose-rates and target cell doses for all particles are not equal; they can vary significantly, in direct contrast to the assumption of dose-equivalency implicit in the use of mass-based media concentrations as metrics of exposure for dose-response assessment. The difference between equivalent nominal media concentration exposures on a μg/mL basis and target cell doses on a particle surface area or number basis can be as high as three to six orders of magnitude. As a consequence, in vitro hazard assessments utilizing mass-based exposure metrics have inherently high errors where particle number or surface areas target cells doses are believed to drive response. The gold standard for particle dosimetry for in vitro nanotoxicology studies should be direct experimental measurement of the cellular content of the studied particle. However, where such measurements are impractical, unfeasible, and before such measurements become common, particle dosimetry models such as ISDD provide a valuable, immediately useful alternative, and eventually, an adjunct to such measurements.

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Polycyclic aromatic hydrocarbons (PAHs) have toxic impacts on humans and ecosystems. One of the most carcinogenic PAHs, benzo(a)pyrene (BaP), is efficiently bound to and transported with atmospheric particles. Laboratory measurements show that particle-bound BaP degrades in a few hours by heterogeneous reaction with ozone, yet field observations indicate BaP persists much longer in the atmosphere, and some previous chemical transport modeling studies have ignored heterogeneous oxidation of BaP to bring model predictions into better agreement with field observations. We attribute this unexplained discrepancy to the shielding of BaP from oxidation by coatings of viscous organic aerosol (OA). Accounting for this OA viscosity-dependent shielding, which varies with temperature and humidity, in a global climate/chemistry model brings model predictions into much better agreement with BaP measurements, and demonstrates stronger long-range transport, greater deposition fluxes, and substantially elevated lung cancer risk from PAHs. Model results indicate that the OA coating is more effective in shielding BaP in the middle/high latitudes compared with the tropics because of differences in OA properties (semisolid when cool/dry vs. liquid-like when warm/humid). Faster chemical degradation of BaP in the tropics leads to higher concentrations of BaP oxidation products over the tropics compared with higher latitudes. This study has profound implications demonstrating that OA strongly modulates the atmospheric persistence of PAHs and their cancer risks.