Cancer development due to fiber-like straight type of multi-walled carbon nanotubes (MWCNTs) has raised concerns for human safety because of its shape similar to asbestos. To set concentrations of MWCNT for a rat carcinogenicity study, we conducted a 13-week whole body inhalation study. F344 male and female rats, 6-week-old at the commencement of the study, were exposed by whole-body inhalation to MWCNT at concentrations of 0, 0.2, 1 and 5 mg/m(3) with a generation and exposure system utilizing the cyclone sieve method. Measured concentrations in the exposure chambers were 0.20 ± 0.02, 1.01 ± 0.11 and 5.02 ± 0.25 mg/m(3) for 13 weeks. The MMAD (GSD) of MWCNT were 1.4-1.6 μm (2.3-3.0), and mean width and length were 94.1-98.0 nm and 5.53-6.19 μm, respectively, for each target concentration. Lung weights were increased 1.2-fold with 1 mg/m(3) and 1.3-fold with 5 mg/m(3) in both sexes compared to the controls. In the bronchoalveolar lavage fluid (BALF) analyses, inflammatory parameters were increased concentration-dependently in both sexes from 0.2 mg/m(3). Granulomatous changes in the lung were induced at 1 and 5 mg/m(3) in females and even at 0.2 mg/m(3) in males. Focal fibrosis of the alveolar wall was observed in both sexes at 1 mg/m(3) or higher. Inflammatory infiltration in the visceral pleural and subpleural areas was induced only at 5 mg/m(3). In conclusion, we determined 0.2 mg/m(3) as the low-observed-adverse-effect level (LOAEL) for respiratory tract toxicity in the present inhalation exposure study of rats.

Inhalation Carcinogenicity and ChronicToxicity of Indium-tin Oxide in Rats and Mice: Kasuke NagaNo, et al. Japan Bioassay Research Center, Japan Industrial Safety and Health Association-Objectives: Carcinogenicity and chronic toxicity of indium-tin oxide (ITO) were examined by inhalation exposure of rats and mice to ITO aerosol. Methods: Fifty mice of both sexes were exposed to ITO at 0 (control), 0.01, 0.03 or 0.1 mg/m 3 for 6 h/day, 5 day/wk for 104 wk, and 50 rats of both sexes were exposed to 0, 0.01 or 0.03 mg/m 3 ITO for the same time period. The repeated exposure of 50 rats of both sexes to 0.1 mg/m 3 ITO was discontinued at the 26th wk, followed by clean air exposure for the remaining 78 wk. Results: In rats, incidences of bronchiolo-alveolar adenomas and carcinomas, bronchiolo-alveolar hyperplasia, alveolar wall fibrosis and thickened pleural wall, alveolar proteinosis and infiltrations of alveolar macrophages and inflammatory cells were significantly increased. Combined incidences of malignant lung tumors in male rats and total lung tumors in male and female rats were significantly increased at exposure to 0.01 mg/m 3 ITO. In mice, no carcinogenic response occurred, but thickened pleural wall, alveolar proteinosis and alveolar macrophage infiltration were induced. Mice were less susceptible to ITO than rats. The lung content of indium was the greatest, followed by the spleen, kidney and liver. Blood indium levels increased dosedependently. Conclusions: There was clear evidence of carcinogenicity of inhaled ITO in male and female rats but not clear evidence in mice, together with occurrence of the chronic pulmonary lesions in both rats and mice. (J Occup Health 2011; 53: 175-187)

Two-and 13-week Inhalation Toxicities of Indium-tin Oxide and Indium Oxide in Rats: Kasuke NagaNo, et al. Japan Bioassay Research Center, Japan Industrial Safety and Health AssociationObjectives: Two-and 13-week inhalation toxicities of indium-tin oxide (ITO) and indium oxide (IO) were characterized for risk assessments of workers exposed to ITO. Methods: F344 rats of both sexes were exposed by inhalation to ITO or IO aerosol for 6 h/day, 5 day/wk for 2 wk at 0, 0.1, 1, 10 or 100 mg/m 3 or 13 wk at 0, 0.1or 1 mg/m 3 . An aerosol generator and inhalation exposure system was constructed. Results: Blood and lung contents of indium were elevated in a dose-related manner in the ITO-and IO-exposed rats. ITO and IO particles were deposited in the lung, mediastinal lymph node and nasal-associated lymphoid tissue. Exposures to ITO and IO induced alveolar proteinosis, infiltrations of alveolar macrophages and inflammatory cells and alveolar epithelial hyperplasia in addition to increased lung weight. ITO affected the lung more severely than IO did. Fibrosis of alveolar wall developed and some of these lesions worsened at the end of the 26-week post-exposure period. Conclusions: Persistent pulmonary lesions including alveolar proteinosis and macrophage infiltration occurred after 2-and 13-week inhalation exposures of rats to ITO and IO. Fibrosis of alveolar wall developed later. These lesions occurred after ITO exposure at the same concentration as the current occupational exposure limit in the USA and at blood indium levels below the biological exposure index in Japan for indium. (J Occup Health 2011; 53: 51-63)

To evaluate pulmonary toxicity of multi-walled carbon nanotubes (MWCNTs), F344 rats of both sexes were exposed by inhalation to 0.2, 1 or 5 mg/m3 MWCNT aerosol for 6 h/day, 5 days/week for 2 weeks using a whole-body exposure system. At the end of the 2-week exposure period, one-half of the rats were necropsied, and at the end of an additional 4-week postexposure period, the remaining rats were necropsied. MWCNTs were deposited in the lungs of all MWCNT-exposed groups and mostly remained in the lungs throughout the 4-week postexposure period. Granulomatous changes in the lung were found in the rats exposed to 5 mg/m3 MWCNTs, and these changes were slightly aggravated at the end of the 4-week postexposure period. In the bronchoalveolar lavage fluid (BALF), the numbers of neutrophils, percentages of bi- and multinucleated alveolar macrophages, levels of ALP activity and concentrations of total protein and albumin were elevated in the rats exposed to 1 and 5 mg/m3 MWCNTs. At the end of the 4-week postexposure period, the values of the BALF parameters tended to remain elevated. In addition, goblet cell hyperplasias in the nasal cavity and nasopharynx were observed in the rats exposed to 1 and 5 mg/m3 MWCNTs, but these lesions had largely regressed by the end of the postexposure period. Based on the histopathological and inflammatory changes, the no-observed-adverse-effect level (NOAEL) for inhalation of MWCNTs for 2 weeks was 0.2 mg/m3.

Because the primary route of human exposure to multi-walled carbon nanotube (MWCNT) is via inhalation, a new dry MWCNT aerosol generation and exposure system for whole-body inhalation exposure using a cyclone and sieve has been developed. The system was tested for operational performance at 0.2, 1 and 5 mg/m(3). Additionally, it was examined whether this system can be employed in animal whole-body inhalation studies by exposing rats to MWCNT aerosol for 6 h at 5 mg/m(3). The system could consistently provide aerosols with a similar particle size distribution and configuration at all the target exposure concentrations. Almost all MWCNTs were fibrous, and the presence of many well-dispersed, nano-sized particles was confirmed. Additionally, the animal study revealed that large amounts of MWCNTs were inhaled into the lung, resulting in an inflammatory response, with increased LDH and albumin levels, and granulomatous change. Therefore, the aerosol generation and exposure system appears useful for MWCNT inhalation studies using rats.