Assessing nanotoxicity in cells in vitro

Abstract

Nanomaterials are commonly defined as particles or fibers of less than 1 micron in diameter. For these reasons, they may be respirable in humans and have the potential, based upon their geometry, composition, size and transport or durability in the body, to cause adverse effects on human health, especially if they are inhaled at high concentrations. Rodent inhalation models to predict the toxicity and pathogenicity of nanomaterials are prohibitive in terms of time and expense. For these reasons, a panel of in vitro assays is described below. These include cell culture assays for cytotoxicity (altered metabolism, decreased growth, lytic or apoptotic cell death), proliferation, genotoxicity, and altered gene expression. The choice of cell type for these assays may be dictated by the procedure or endpoint selected. Most of these assays have been standardized in our laboratory using pathogenic minerals (asbestos, silica) and nonpathogenic particles (fine titanium dioxide or glass beads) as negative controls. The results of these in vitro assays should predict whether testing of selected nanomaterials should be pursued in animal inhalation models that simulate physiologic exposure to inhaled nanomaterials. Conversely, intrathoracic or intrapleural injection of nanomaterials into rodents can be misleading as they bypass normal clearance mechanisms, and nonpathogenic fibers and particles can test positively in these assays.

Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro–in vivo gap

“…In vitro assays are mainly the first to be conducted during a toxicological evaluation 78 . Most in vitro studies are conducted in classical 2D monocultures of cancer or long-lived cell lines although the use of stem cells or primary cells is steadily increasing.…”

“…The selection of a relevant cell type and source generally depends on the expected in vivo target organ and application of the NP 79 . In the vast majority of the studies the cells are exposed to the NP dispersion during a single incubation period ranging from 3 up to 48 hours 78,80,81 .…”

“…Furthermore the possibility of upscaling and automation of the execution, detection and data processing make these assays highly convenient with regard to a future high content screening approach for every newly synthetized NP 83,84 . Another popular approach is staining the entire cell or specific cellular components with cellular dyes, fluorescently labeled antibodies (AB) or molecular probes, which will interact with a specific biomolecule 40,78,85 . The samples can on the one hand be analyzed using a plate reader, but if information is requested on the variation between different cell types or status (e.g.…”

“…The use of such stainings in combination with microscopy however mostly generates qualitative or semi-quantitative data. In order to generate quantitative data, other easily quantifiable assays are required as for example the comet assay, PCR or microarrays to evaluate DNA damage 78 .…”

DNA gel particles: An overview

“…Currently, there are no specific testing requirements for nanotechnology products, and therefore, researchers took liberal approaches to studying toxicity [102][103]. Moreover, it is worth noting that, because of the expense of animal testing in toxicology and pressure from both the general public and government to develop alternatives to in vivo testing, in vitro cell-based models may be more attractive for preliminary testing of nanomaterials [104]. The prediction of toxicity is difficult, but cytotoxicity screening, which is routinely used in drug screening, gives a good indication of potential adverse effects in cells.…”