Carbon black nanoparticle (CBNP) applications in high doses have been shown to be harmful to the lung. It is postulated that even small, environmentally relevant concentrations induce changes on lung homeostasis. The present study determined the impact of low-dose single and multiple CBNP (Printex 90) applications on mouse alveolar cell metabolism, especially inflammatory and oxidative stress parameters. Nanoparticles were administered to mice by a single or 8 oropharyngeal aspirations at wk 1, 2, 3, 5, 7, 9, 11, and 12 using 7 μg Printex 90, 7 μg DQ12 quartz (positive control), with water vehicle and saline as negative controls. After 2 d or 3 mo lung function was analyzed. Further lung histology, bronchoalveolar lavage fluid (BALF) parameters, and mRNA expression of cytokines and antioxidants enzymes in type II pneumocytes were measured on d 3 or after 3 mo. Single low-dose Printex 90 application induced no marked alterations in lung functions or BALF phospholipid levels but significant decrease in superoxide dismutase 2 and numerically elevated glutathione peroxidase 3 mRNA expression levels in type II pneumocytes. Multiple CBNP applications produced reduced lung function, collagen accumulation, elevated phospholipid levels in BALF, and a massive infiltration of macrophages. Type II pneumocyte mRNA expression of antioxidative enzymes remained unchanged throughout the subchronic experiment, but showed a significant decrease in interleukin (IL)-6Rα mRNA expression. This study demonstrates that an environmentally relevant CBNP concentration induced an acute inflammatory response, an effect that is exacerbated throughout the subchronic duration.

Inhalation of nitrogen and reactive oxygen species (ROS) is known to induce lung inflammation, which is prevented by enzymatic and nonenzymatic antioxidant systems. These agents form nitrated allergens that were shown to enhance allergenicity. The aim of this study was to examine the influence of nitrated proteins on inflammation and antioxidant status of the lung. Ovalbumin (OVA) in nitrated form (nOVA) was intraperitoneally (ip) injected in mice for sensitization and in nitrated or unmodified form for challenge to induce allergic bronchial inflammation. To study the allergen potential of unrelated protein and verify cross-reactivity, nitrated and unmodified keyhole limpet hemocyanin (nKLH, KLH) was used for challenge. Challenge with OVA or nOVA reduced lung function and increased eosinophilia and protein content in bronchoalveolar lavage fluid (BALF). Challenge with nitrated or native OVA or KLH elevated glutathione (GSH) ratio in type II pneumocytes. Reduced mRNA expression of glutathione peroxidase (GPX) 3, glutathione reductase (GR), superoxide dismutase (SOD) 2, and catalase (CAT) was most prominent after challenge with nitrated OVA and nitrated KLH, respectively. Challenge with nOVA enhanced SOD1 mRNA reduction. Immunostaining of GPX 3 and SOD2 increased after challenge with OVA or nOVA, while reactivity of GR and reactivity of SOD2 were reduced after challenge with KLH or nKLH. SOD1 immunostaining was diminished after challenge with nonnitrated OVA or KLH. CAT immunoreaction was similar in all groups. Nitrated proteins without allergenic potential triggered mRNA reduction of antioxidants in type II cells after sensitization with a nitrated allergen but did not induce bronchial inflammation.

Inhalation uptake of carbon black nanoparticles (CBNP) bears the risk of morphological and functional lung impairment attributed to the highly reactive particle surface area. Chemical particle surface modifications might affect particle-cell interactions; however, thus far these alterations have not been determined. This is the first in vivo study comparing particle-induced acute lung injury using Printex(®)90 (Pr90, 7 µg), Printex®90 covered by benzo[a]pyrene or 9-nitroanthracene (BaP-Pr90, NA-Pr90, 7 µg, 15% BaP or NA by weight), and acetylene carbon black (CB) with polycyclic aromatic hydrocarbons (PAH-AB, 7 µg, 20% PAH by weight). All particles were suspended in distilled water with bovine serum albumin (BSA). In addition, the influence of suspension media was tested using Printex®90 suspended without BSA (Pr90(-BSA), 7 µg). Quartz (DQ12, 7 µg), 70 µl saline (NaCl), and distilled water with or without BSA (H2O(+/-BSA)) were used as reference and controls. It was postulated that CBNP surface modifications trigger pulmonary responses. After oropharyngeal particle aspiration, lung functions were measured 2 d postexposure, followed by lung preparation for histological or bronchoalveolar lavage fluid (BALF) examinations and type II pneumocyte isolation on d 3. Head-out body plethysmography revealed reduced flow rates induced by PAH-AB. Examinations of BALF demonstrated reduced influx of macrophages after exposure to Pr90(-BSA) and decreased lymphocyte levels after Pr90(+BSA) or BaP-Pr90 treatment. Further, CBNP induced changes in mRNA expressions (surfactant proteins) in type II pneumocytes. These findings indicate that CBNP surface area and media modulate interactions between NP and lung cells in short-term experiments.

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