At 6-month post-exposure, significant changes were not observed in the group exposed to 0.2 mg/kg MWCNTs. In the group exposed to 1 mg/kg MWCNTs, deposition of the MWCNTs and macrophage accumulation, of which some of them were granulomatous, were observed in the alveoli and interstitium until 6-month post-exposure, although they were minimal changes. Studies have reported that pulmonary fibrosis is induced due to exposure to SWCNTs or MWCNTs (Muller et al., 2005 and Shvedova et al., 2008a); however, pulmonary fibrosis

was not observed in any of the groups in this study. Light microscopy and TEM observations revealed that the MWCNTs deposited in the lungs were phagocytosed by alveolar macrophages and were sequentially accumulated in the alveoli. MWCNT translocation or penetration to the pleural was not observed. Furthermore, based on the 400 TEM images, it was shown that all the MWCNTs were located in the alveolar macrophages or Ruxolitinib phagocytosed by macrophages in the interstitial tissues, and individual MWCNTs were not presented in the cells of the interstitial tissue. In contrast, inflammatory responses were observed in the lungs and lung-associated lymph nodes in the group exposed to 5 mg/kg crystalline silica, where BALF inflammatory cells, LDH, TP, IL-1β, and IL-2 levels were significantly increased after the instillation exposure,

and these changes CTLA-4 inhibiton were the most severe at 6-month post-exposure. Furthermore, lung weights were significantly increased at 3- and 6-month post-exposure. Histopathological evaluation revealed that although short-term inflammatory responses were weak, the inflammatory responses were much stronger at 6-month post-exposure. Consequently, crystalline silica particles produced continuous inflammation with a 5 mg/kg dose of intratracheal instillation. These pulmonary responses

were qualitatively and quantitatively different from the responses observed for MWCNTs instillation exposure. The relationship of the dose of MWCNTs instilled into the lungs in this study and exposure levels of aerosolized MWCNTs to humans during the handling of CNTs in the work place is discussed below. The pulmonary deposition amount Florfenicol of MWCNTs in this study was considered to be almost 100% of the instilled dose of the MWCNTs (i.e., 0.04, 0.2, and 1.0 mg/kg). By measuring the BET surface area of the MWCNT samples, the doses can be expressed in terms of the CNT surface area dose, which are 0.0009, 0.1146, and 0.023 m2/kg, for doses of 0.04, 0.2, and 1.0 mg/kg, respectively. Based on the density of the MWCNT samples reported by the manufacturer (2.1 g/cm3) and assuming that the tube diameter and length are uniform (60 nm and 1.5 μm, respectively), and that all tubes are individually dispersed in the suspension, the doses can also be expressed in terms of tube numbers, which are 9.4 × 109, 4.7 × 1010, and 2.4 × 1011 tubes/kg, for dosed of 0.04, 0.2, and 1.0 mg/kg, respectively.