Byeong-Woo Lee

Comparison of pulmonary inflammatory responses following intratracheal instillation and inhalation of nanoparticles

Abstract In order to examine whether intratracheal instillation studies can be useful for determining the harmful effect of nanoparticles, we performed inhalation and intratracheal instillation studies using samples of the same nanoparticles. Nickel oxide nanoparticles (NiO) and titanium dioxide nanoparticles (TiO2) were used as chemicals with high and low toxicities, respectively. In the intratracheal instillation study, rats were exposed to 0.2 or 1 mg of NiO or TiO2. Cell analysis and chemokines in bronchoalveolar lavage fluid (BALF) were analyzed from 3 days to 6 months following the single intratracheal instillation. In the inhalation study, rats were exposed to inhaled NiO or TiO2 (1.65, 1.84 mg/m3, respectively) for 4 weeks. The same endpoints were examined from 3 days to 3 months after the end of exposure. Inhalation of NiO induced an increase in the number of neutrophils in BALF and concentrations of cytokine-induced neutrophil chemoattractant (CINC)-1, CINC-2 and heme oxygenase (HO)-1. Intratracheal instillation of NiO induced persistent inflammation and upregulation of these cytokines was observed in the rats. However, inhalation of TiO2 did not induce pulmonary inflammation, and intratracheal instillation of TiO2 transiently induced an increase in the number of neutrophils in BALF and the concentrations of CINC-1, CINC-2 and HO-1. Taken together, a difference in pulmonary inflammation was observed between the high and low toxicity nanomaterials in the intratracheal instillation studies, as in the inhalation studies, suggesting that intratracheal instillation studies may be useful for ranking the harmful effects of nanoparticles.

Comparison of the Pulmonary Oxidative Stress Caused by Intratracheal Instillation and Inhalation of NiO Nanoparticles when Equivalent Amounts of NiO Are Retained in the Lung

NiO nanoparticles were administered to rat lungs via intratracheal instillation or inhalation. During pulmonary toxicity caused by NiO nanoparticles, the induction of oxidative stress is a major factor. Both intratracheal instillation and inhalation of NiO nanoparticles induced pulmonary oxidative stress. The oxidative stress response protein, heme oxygenase-1 (HO-1), was induced by the administration of NiO nanoparticles at both the protein and gene expression level. Additionally, certain oxidative-stress markers in the lung, such as 8-iso-prostaglandin F2α, thioredoxin, and inducible nitric oxide synthase were increased. Furthermore, the concentration of myeloperoxidase (MPO) in the lung was also increased by the administration of NiO nanoparticles. When the amount of NiO in the lung is similar, the responses against pulmonary oxidative stress of intratracheal instillation and inhalation are also similar. However, the state of pulmonary oxidative stress in the early phase was different between intratracheal instillation and inhalation, even if the amount of NiO in the lung was similar. Inhalation causes milder oxidative stress than that caused by intratracheal instillation. On evaluation of the nanoparticle-induced pulmonary oxidative stress in the early phase, we should understand the different states of oxidative stress induced by intratracheal instillation and inhalation.

Analysis of pulmonary surfactant in rat lungs after inhalation of nanomaterials: Fullerenes, nickel oxide and multi-walled carbon nanotubes

Abstract The health risks of inhalation exposure to engineered nanomaterials in the workplace are a major concern in recent years, and hazard assessments of these materials are being conducted. The pulmonary surfactant of lung alveoli is the first biological entity to have contact with airborne nanomaterials in inhaled air. In this study, we retrospectively evaluated the pulmonary surfactant components of rat lungs after a 4-week inhalation exposure to three different nanomaterials: fullerenes, nickel oxide (NiO) nanoparticles and multi-walled carbon nanotubes (MWCNT), with similar levels of average aerosol concentration (0.13–0.37 mg/m3). Bronchoalveolar lavage fluid (BALF) of the rat lungs stored after previous inhalation studies was analyzed, focusing on total protein and the surfactant components, such as phospholipids and surfactant-specific SP-D (surfactant protein D) and the BALF surface tension, which is affected by SP-B and SP-C. Compared with a control group, significant changes in the BALF surface tension and the concentrations of phospholipids, total protein and SP-D were observed in rats exposed to NiO nanoparticles, but not in those exposed to fullerenes. Surface tension and the levels of surfactant phospholipids and proteins were also significantly different in rats exposed to MWCNTs. The concentrations of phospholipids, total protein and SP-D and BALF surface tension were correlated significantly with the polymorphonuclear neutrophil counts in the BALF. These results suggest that pulmonary surfactant components can be used as measures of lung inflammation.

Analysis of pulmonary surfactant in rat lungs after intratracheal instillation of short and long multi-walled carbon nanotubes.

Abstract Multi-walled carbon nanotubes (MWCNTs) are interesting new materials, but there is some concern about their harmfulness due to their fibrous nature. To determine the difference in the biological effects of MWCMTs by fiber length, we prepared two MWCNT samples from one bulk sample. One consisted of cut up short fibers (Short; average length = 0.94 µm) and the other was just dispersed (Long; average length = 3.4 µm). The samples were administered to male Wistar rats by intratracheal instillation at doses of 0.2 mg and 1 mg/animal (Short) and 0.2 mg and 0.6 mg/animal (Long). The animals were sacrificed at time points from 3 d to 12 months after administration. Bronchoalveolar lavage fluid (BALF) was taken from the lungs and pathological specimens were prepared. The concentrations of phospholipids, total protein and surfactant protein D (SP-D) in the pulmonary surfactant of the BALF were determined, the surface tension of BALF was measured, and the inflammation score was determined by the point-counting method to assess pulmonary tissue inflammation. The present study suggests that inflammatory response in the lung was slightly higher for long MWCNTs than for short MWCNTs when compared at the same mass dose. The correlation between pulmonary surfactant components and BALF surface tension was also evaluated. The Spearman’s rank correlation coefficients obtained for the phospholipid, total protein and SP-D concentrations were −0.068 (p = 0.605), −0.360 (p = 0.005) and −0.673 (p = 0.000), respectively. Surface tension, measured by a simple method, should be reflected in the change of a surfactant protein, such as SP-D.

Assessment of Pulmonary Toxicity Induced by Inhaled Toner with External Additives

We investigated the harmful effects of exposure to a toner with external additives by a long-term inhalation study using rats, examining pulmonary inflammation, oxidative stress, and histopathological changes in the lung. Wistar rats were exposed to a well-dispersed toner (mean of MMAD: 2.1 μm) at three mass concentrations of 1, 4, and 16 mg/m3 for 22.5 months, and the rats were sacrificed after 6 months, 12 months, and 22.5 months of exposure. The low and medium concentrations did not induce statistically significant pulmonary inflammation, but the high concentration did, and, in addition, a histopathological examination showed fibrosis in the lung. Although lung tumor was observed in one sample of high exposure for 22.5 months, the cause was not statistically significant. On the other hand, a persistent increase in 8-OHdG was observed in the high exposure group, indicating that DNA damage by oxidative stress with persistent inflammation leads to the formation of tumorigenesis. The results of our studies show that toners with external additives lead to pulmonary inflammation, oxidative stress, and fibrosis only at lung burdens beyond overload. These data suggest that toners with external additives may have low toxicity in the lung.

Biopersistence of NiO and TiO2 Nanoparticles Following Intratracheal Instillation and Inhalation

The hazards of various types of nanoparticles with high functionality have not been fully assessed. We investigated the usefulness of biopersistence as a hazard indicator of nanoparticles by performing inhalation and intratracheal instillation studies and comparing the biopersistence of two nanoparticles with different toxicities: NiO and TiO2 nanoparticles with high and low toxicity among nanoparticles, respectively. In the 4-week inhalation studies, the average exposure concentrations were 0.32 and 1.65 mg/m3 for NiO, and 0.50 and 1.84 mg/m3 for TiO2. In the instillation studies, 0.2 and 1.0 mg of NiO nanoparticles and 0.2, 0.36, and 1.0 mg of TiO2 were dispersed in 0.4 mL water and instilled to rats. After the exposure, the lung burden in each of five rats was determined by Inductively Coupled Plasma-Atomic Emission Spectrometer (ICP-AES) from 3 days to 3 months for inhalation studies and to 6 months for instillation studies. In both the inhalation and instillation studies, NiO nanoparticles persisted for longer in the lung compared with TiO2 nanoparticles, and the calculated biological half times (BHTs) of the NiO nanoparticles was longer than that of the TiO2 nanoparticles. Biopersistence also correlated with histopathological changes, inflammatory response, and other biomarkers in bronchoalveolar lavage fluid (BALF) after the exposure to nanoparticles. These results suggested that the biopersistence is a good indicator of the hazards of nanoparticles.