Although amorphous silica nanoparticles are widely used in the production of food products (e.g., as anticaking agents), there is little information available about their absorption and biological effects after oral exposure. Here, we examined the in vitro intestinal absorption and in vivo biological effects in mice of orally administered amorphous silica particles with diameters of 70, 300, and 1,000 nm (nSP70, mSP300, and mSP1000, respectively) and of nSP70 that had been surface-modified with carboxyl or amine groups (nSP70-C and nSP70-N, respectively). Analysis of intestinal absorption by means of the everted gut sac method combined with an inductively coupled plasma optical emission spectrometer showed that the intestinal absorption of nSP70-C was significantly greater than that of nSP70. The absorption of nSP70-N tended to be greater than that of nSP70; however, the results were not statistically significant. Our results indicate that silica nanoparticles can be absorbed through the intestine and that particle diameter and surface properties are major determinants of the degree of absorption. We also examined the biological effects of the silica particles after 28-day oral exposure in mice. Hematological, histopathological, and biochemical analyses showed no significant differences between control mice and mice treated with the silica particles, suggesting that the silica nanoparticles evaluated in this study are safe for use in food production.

The application of nanotechnology in the health care setting has many potential benefits; however, our understanding of the interactions between nanoparticles and our immune system remains incomplete. Although many of the biological effects of nanoparticles are negatively correlated with particle size, some are clearly size specific and the mechanisms underlying these size-specific biological effects remain unknown. Here, we examined the pro-inflammatory effects of silica particles in THP-1 cells with respect to particle size; a large overall size range with narrow intervals between particle diameters (particle diameter: 10, 30, 50, 70, 100, 300, and 1,000 nm) was used. Secretion of the pro-inflammatory cytokines interleukin (IL)-1β and tumor necrosis factor (TNF)-α induced by exposure to the silica particles had a bell-shaped distribution, where the maximal secretion was induced by silica nanoparticles with a diameter of 50 nm and particles with smaller or larger diameters had progressively less effect. We found that blockade of IL-1β secretion markedly inhibited TNF-α secretion, suggesting that IL-1β is upstream of TNF-α in the inflammatory cascade induced by exposure to silica particles, and that the induction of IL-1β secretion was dependent on both the NLRP3 inflammasome and on uptake of the silica particles into the cells via endocytosis. However, a quantitative analysis of silica particle uptake showed that IL-1β secretion was not correlated with the amount of silica particles taken up by the cells. Further investigation revealed that the induction of IL-1β secretion and uptake of silica nanoparticles with diameters of 50 or 100 nm, but not of 10 or 1,000 nm, was dependent on scavenger receptor (SR) A1. In addition, of the silica particles examined, only those with a diameter of 50 nm induced strong IL-1β secretion via activation of Mer receptor tyrosine kinase, a signal mediator of SR A1. Together, our results suggest that the SR A1-mediated pro-inflammatory response is dependent on ligand size and that both SR A1-mediated endocytosis and receptor-mediated signaling are required to produce the maximal pro-inflammatory response to exposure to silica particles.

Vaccine adjuvants that can induce not only antigen-specific antibody responses but also Th1-type immune responses and CD8+ cytotoxic T lymphocyte responses are needed for the development of vaccines against infectious diseases and cancer. Of many available adjuvants, oligodeoxynucleotides (ODNs) with unmethylated cytosine-phosphate-guanine (CpG) motifs are the most promising for inducing the necessary immune responses, and these adjuvants are currently under clinical trials in humans. However, the development of novel delivery vehicles that enhance the adjuvant effects of CpG ODNs, subsequently increasing the production of cytokines such as type-I interferons (IFNs), is highly desirable. In this study, we demonstrate the potential of pH-responsive biodegradable carbonate apatite (CA) nanoparticles as CpG ODN delivery vehicles that can enhance the production of type-I IFNs (such as IFN-α) relative to that induced by CpG ODNs and can augment the adjuvant effects of CpG ODNs in vivo. In contrast to CpG ODNs, CA nanoparticles containing CpG ODNs (designated CA-CpG) induced significant IFN-α production by mouse dendritic cells and human peripheral blood mononuclear cells in vitro; and production of interleukin-12, and IFN-γ was higher in CA-CpG-treated groups than in CpG ODNs groups. In addition, treatment with CA-CpG resulted in higher cytokine production in draining lymph nodes than did treatment with CpG ODNs in vivo. Furthermore, vaccination with CA-CpG plus an antigen, such as ovalbumin or influenza virus hemagglutinin, resulted in higher antigen-specific antibody responses and CD8+ cytotoxic T lymphocyte responses in vivo, in an interleukin-12- and type-I IFN-dependent manner, than did vaccination with the antigen plus CpG ODNs; in addition, the efficacy of the vaccine against influenza virus was higher with CA-CpG as the adjuvant than with CpG ODNs as the adjuvant. These data show the potential of CA nanoparticles to serve as CpG ODN delivery vehicles that increase the production of cytokines, especially IFN-α, induced by CpG ODNs and thus augment the efficacy of CpG ODNs as adjuvants. We expect that the strategy reported herein will facilitate the design and development of novel adjuvant delivery vehicles for vaccines.