Inhalation exposure study of titanium dioxide nanoparticles with a primary particle size of 2 to 5 nm

Abstract

Background: Nanotechnology offers great promise in many industrial applications. However, little is known about the health effects of manufactured nanoparticles, the building blocks of nanomaterials.

Objectives: Titanium dioxide (TiO(2)) nanoparticles with a primary size of 2-5 nm have not been studied previously in inhalation exposure models and represent some of the smallest manufactured nanoparticles. The purpose of this study was to assess the toxicity of these nanoparticles using a murine model of lung inflammation and injury.

Materials and methods: The properties of TiO(2) nanoparticles as well as the characteristics of aerosols of these particles were evaluated. Mice were exposed to TiO(2) nanoparticles in a whole-body exposure chamber acutely (4 hr) or subacutely (4 hr/day for 10 days). Toxicity in exposed mice was assessed by enumeration of total and differential cells, determination of total protein, lactate dehydrogenase (LDH) activity and inflammatory cytokines in bronchoalveolar lavage (BAL) fluid. Lungs were also evaluated for histopathologic changes

Results: Mice exposed acutely to 0.77 or 7.22 mg/m(3) nanoparticles demonstrated minimal lung toxicity or inflammation. Mice exposed subacutely (8.88 mg/m(3)) and necropsied immediately and at week 1 or 2 postexposure had higher counts of total cells and alveolar macrophages in the BAL fluid compared with sentinels. However, mice recovered by week 3 postexposure. Other indicators were negative.

Conclusions: Mice subacutely exposed to 2-5 nm TiO(2) nanoparticles showed a significant but moderate inflammatory response among animals at week 0, 1, or 2 after exposure that resolved by week 3 postexposure.

Figure 1

Small whole-body exposure chamber used in these studies for nanoparticle inhalation exposure studies. An aerosol-laden flow stream is generated with a nebulizer. After passing through a dryer the flow stream is sent into the exposure chamber. Nanoparticle concentrations and size distributions are measured using gravimetrical analysis and the SMPS, respectively. A TEM stub placed inside of the exposure chamber is also used for characterization. See ”Materials and Methods” for further details.

Figure 2

TEM images of dispersed (A) and aggregated (B) TiO₂ nanoparticles. Dispersed nanoparticles show a primary nanoparticle size between 2 and 5 nm. For the generated aerosol, the TiO₂ particles aggregate to form larger particles as shown in B.

Figure 3

(A) XPS spectrum in the O(1s) region show the presence of both O atoms and O–H groups on the surface of the TiO₂ nanoparticles. (B) The ATR-FTIR spectrum of TiO₂ nanoparticles under ambient conditions. The absorption bands in the spectrum are associated with the bending, δ(H₂O), and stretching, ν(H₂O), modes of adsorbed water at 1,645 and 3,400 cm⁻¹, respectively. The absorption band below 1,000 cm⁻¹ is due to TiO₂ lattice vibrations.

Figure 4

Number of macrophages, neutrophils and lymphocytes in BAL fluid among acutely (A) and subacutely (B) exposed animals. Values are expressed as mean ± SE. *Significantly different from control group, p < 0.05 (t-test for equal and unequal variances).

Figure 5

Dark field micrographs of lung tissue with H&E staining (A,B) and alveolar macrophages prepared by cytospinning and H&E staining (C,D). (A,C) Sentinels and (B,D) animals subacutely exposed to TiO₂ nanoparticles with a primary particle size of 2–5 nm and necropsied immediately after the last exposure. Arrows point to TiO₂ nanoparticle-laden macrophages.

Figure 6

Dark field micrographs of alveolar macrophages prepared by cytospinning and H&E staining from mice exposed subacutely to TiO₂ nanoparticles and necropsied at weeks 0 (A), 1 (B), 2 (C), and 3 (D) postexposure.