Oxidative stress-mediated inhibition of intestinal epithelial cell proliferation by silver nanoparticles.

Given the increasing use of silver nanoparticles (Ag NP) by the food and food packaging industries, this study investigated potential consequences of Ag NP ingestion in intestinal epithelial C2BBe1 cells. Treatment of proliferating cells (<10,000 cells/cm(2)) with 0.25 µg/cm(2) (1.25 µg/mL) of 23 nm Ag NP for 24 h induced 15% necrotic cell death and an 80% reduction in metabolic activity and decreased the GSH/GSSG ratio, indicating oxidative stress. G2/M phase cell cycle arrest and complete inhibition of cell proliferation was also induced by Ag NP treatment. Simulated in vitro digestion of Ag NP prior to cell exposure required the use of slightly higher doses to induce the same toxicity, likely due to slower Ag dissolution. Treatment of cells with silica, titania, and ZnO NP partially inhibited cell proliferation, but inhibition at low doses was unique to Ag NP. These data suggest that Ag NP induces oxidative stress, cell cycle arrest, and the inhibition of cell proliferation. However, toxicity and induction of oxidative stress were not observed in confluent cells (>100,000 cells/cm(2)) treated with 10 µg/cm(2) (40-50 µg/mL) Ag NP, indicating that these cells are less sensitive to Ag NP.

Uptake of bright fluorophore core-silica shell nanoparticles by biological systems.

Nanoparticles are used in a variety of consumer applications. Silica nanoparticles in particular are common, including as a component of foods. There are concerns that ingested nano-silica particles can cross the intestinal epithelium, enter the circulation, and accumulate in tissues and organs. Thus, tracking these particles is of interest, and fluorescence spectroscopic methods are well-suited for this purpose. However, nanosilica is not fluorescent. In this article, we focus on core-silica shell nanoparticles, using fluorescent Rhodamine 6G, Rhodamine 800, or CdSe/CdS/ZnS quantum dots as the core. These stable fluorophore/silica nanoparticles had surface characteristics similar to those of commercial silica particles. Thus, they were used as model particles to examine internalization by cultured cells, including an epithelial cell line relevant to the gastrointestinal tract. Finally, these particles were administered to mice by gavage, and their presence in various organs, including stomach, small intestine, cecum, colon, kidney, lung, brain, and spleen, was examined. By combining confocal fluorescence microscopy with inductively coupled plasma mass spectrometry, the presence of nanoparticles, rather than their dissolved form, was established in liver tissues.

Γ-tubulin controls neuronal microtubule polarity independently of Golgi outposts.

Neurons have highly polarized arrangements of microtubules, but it is incompletely understood how microtubule polarity is controlled in either axons or dendrites. To explore whether microtubule nucleation by γ-tubulin might contribute to polarity, we analyzed neuronal microtubules in Drosophila containing gain- or loss-of-function alleles of γ-tubulin. Both increased and decreased activity of γ-tubulin, the core microtubule nucleation protein, altered microtubule polarity in axons and dendrites, suggesting a close link between regulation of nucleation and polarity. To test whether nucleation might locally regulate polarity in axons and dendrites, we examined the distribution of γ-tubulin. Consistent with local nucleation, tagged and endogenous γ-tubulins were found in specific positions in dendrites and axons. Because the Golgi complex can house nucleation sites, we explored whether microtubule nucleation might occur at dendritic Golgi outposts. However, distinct Golgi outposts were not present in all dendrites that required regulated nucleation for polarity. Moreover, when we dragged the Golgi out of dendrites with an activated kinesin, γ-tubulin remained in dendrites. We conclude that regulated microtubule nucleation controls neuronal microtubule polarity but that the Golgi complex is not directly involved in housing nucleation sites.

Minimal intestinal epithelial cell toxicity in response to short- and long-term food-relevant inorganic nanoparticle exposure.

Toxicity of commercial nanoparticles of titania, silica, and zinc oxides is being investigated in this in vitro study. Particles of these compositions are found in many food items, and thus this study is directed toward particle behavior in simulated digestion media and their interaction with intestinal epithelial cell line C2BBe1, a clone of Caco-2 cells, originally isolated from a human colon cancer. Even though the primary particle size of all three particles was below 50 nm, the particles appeared as aggregates in culture media with a negatively charged surface. In the presence of pepsin (pH 2), the charge on the titania became positive, and silica was almost neutral and aggregated extensively, whereas ZnO dissolved. For silica and titania, treatment with simulated intestinal digestive solution led to a strongly negatively charged surface and particle sizes approaching values similar to those in media. On the basis of infrared spectroscopy, we concluded that the surface of silica and titania was covered with bile salts/proteins after this treatment. Transmission electron microscopy indicated that the C2BBe1 cells internalized all three particles. Toxicity assays included investigation of necrosis, apoptosis, membrane damage, and mitochondrial activity. Titania and SiO2 particles suspended in media at loading levels of 10 µg/cm² exhibited no toxicity. With ZnO at the same loading level, mild toxicity was observed based only on the LDH assay and decrease of mitochondrial activity and not necrosis or apoptosis. Titania particles exposed to the simulated digestion media exhibited mild toxicity based on decrease of mitochondrial activity, likely due to transport of toxic bile salts via adsorption on the particle surface.