The evaluation of the role of physicochemical properties in the toxicity of nanoparticles is important for the understanding of toxicity mechanisms and for controlling the behavior of nanoparticles. The surface charge of nanoparticles is suggested as one of the key parameters which decide their biological impact. In this study, we synthesized fluorophore-conjugated polystyrene nanoparticles (F-PLNPs), with seven different types of surface functional groups that were all based on an identical core, to evaluate the role of surface charge in the cellular uptake of nanoparticles. Phagocytic differentiated THP-1 cells or non-phagocytic A549 cells were incubated with F-PLNP for 4 h, and their cellular uptake was quantified by fluorescence intensity and confocal microscopy. The amount of internalized F-PLNPs showed a good positive correlation with the zeta potential of F-PLNPs in both cell lines (Pearson’s r = 0.7021 and 0.7852 for zeta potential vs. cellular uptake in THP-1 cells and nonphagocytic A549 cells, respectively). This result implies that surface charge is the major parameter determining cellular uptake efficiency, although other factors such as aggregation/agglomeration, protein corona formation, and compositional elements can also influence the cellular uptake partly or indirectly.

Background: The quantification of nanomaterials accumulated in various organs is crucial in studying their toxicity and toxicokinetics. However, some types of nanomaterials, including carbon nanomaterials (CNMs), are difficult to quantify in a biological matrix. Therefore, developing improved methodologies for quantification of CNMs in vital organs is instrumental in their continued modification and application. Results: In this study, carbon black, nanodiamond, multi-walled carbon nanotube, carbon nanofiber, and graphene nanoplatelet were assembled and used as a panel of CNMs. All CNMs showed significant absorbance at 750 nm, while their bio-components showed minimal absorbance at this wavelength. Quantification of CNMs using their absorbance at 750 nm was shown to have more than 94% accuracy in all of the studied materials. Incubating proteinase K (PK) for 2 days with a mixture of lung tissue homogenates and CNMs showed an average recovery rate over 90%. The utility of this method was confirmed in a murine pharyngeal aspiration model using CNMs at 30 μg/mouse. Conclusions: We developed an improved lung burden assay for CNMs with an accuracy > 94% and a recovery rate > 90% using PK digestion and UV-Vis spectrophotometry. This method can be applied to any nanomaterial with sufficient absorbance in the near-infrared band and can differentiate nanomaterials from elements in the body, as well as the soluble fraction of the nanomaterial. Furthermore, a combination of PK digestion and other instrumental analysis specific to the nanomaterial can be applied to organ burden analysis.

Multi-walled carbon nanotubes (MWCNTs) have variable metal impurities, but little is known about the impact of soluble metal impurities on the toxicity of MWCNTs. Here, we evaluated the role of soluble metal impurities to the acute inflammogenic potential of MWCNTs, using five types of high purity MWCNTs (>95%). MWCNTs and their soluble fractions collected at 24 h after incubation in phosphate-buffered saline showed diverse metal impurities with variable concentrations. The fiber-free soluble fractions produced variable levels of reactive oxygen species (ROS), and the iron level was the key determinant for ROS production. The acute inflammation at 24 h after intratracheal instillation of MWCNTs to rats at 0.19, 0.63, and 1.91 mg MWCNT/kg body weight (bw) or fiber-free supernatants from MWCNT suspensions at 1.91 and 7.64 mg MWCNT/kg bw showed that the number of granulocytes, a marker for acute inflammation, was significantly increased with a good dose-dependency. The correlation study showed that neither the levels of iron nor the ROS generation potential of the soluble fractions showed any correlations with the inflammogenic potential. However, the total concentration of transition metals in the soluble fractions showed a good correlation with the acute lung inflammogenic potential. These results implied that metal impurities, especially transitional metals, can contribute to the acute inflammogenic potential of MWCNTs, although the major parameter for the toxicity of MWCNTs is size and shape.

Background: Nanodiamonds (NDs) have gained a rapidly growing interest in biomedical applications; however, little is known regarding their biokinetics owing to difficulties in measurements and limited synthesis/purification technologies. In this study, we investigated the distribution kinetics of detonation-synthesized NDs in mice via intravenous injection to evaluate the parameters that determine the behavior of the particles. We prepared two distinctive NDs that controlled the sp³/sp² carbon ratio and particle size by coating them with serum proteins. The four control samples were intravenously injected into mice, and tissue distribution and clearance were evaluated at 30 min and 1, 7, and 28 days post-injection. Results: The sp³/sp² carbon ratio showed no correlation with the organ distribution of the NDs. However, hydrodynamic size showed an excellent correlation with organ distribution levels: a negative correlation in the liver and positive correlations in the spleen and lungs. Furthermore, the deposition levels of NDs in the lung suggest that particles smaller than 300 nm could avoid lung deposition. Finally, a similar organ distribution pattern was observed in mice injected with carbon black nanoparticles controlled hydrodynamic size. Conclusions: In conclusion, the tissue distribution of NDs is modulated not by the sp³/sp² carbon ratio but by the hydrodynamic size, which can provide helpful information for targeting the tissue of NDs. Furthermore, the organ distribution pattern of the NDs may not be specific to NDs but also can apply to other nanoparticles, such as carbon black.

Pulmonary exposure to sodium fluoride (NaF) may occur in certain consumer-product and occupational settings, but its pulmonary toxicity and systemic biodistribution remain insufficiently characterized. This study investigated acute pulmonary inflammatory responses and biodistribution kinetics in rats following a single airway exposure to NaF. Non-radioactive NaF was administered in 7-week-old male Sprague-Dawley rats by intratracheal instillation at 0.75, 1.5, or 3.0 mg/rat, and pulmonary responses were evaluated at 1, 7, and 28 days using bronchoalveolar lavage fluid analysis and histopathology. Biodistribution was assessed using positron emission tomography/computed tomography (PET/CT) and organ gamma counting after administration of fluorine-18 radiolabeled sodium fluoride ([¹⁸F]NaF). NaF induced acute pulmonary inflammation characterized by increased neutrophils and eosinophils, elevated lactate dehydrogenase activity, and increased levels of tumor necrosis factor-alpha and eotaxin. These inflammatory responses gradually resolved by 7 and 28 days after exposure. PET/CT analysis showed rapid pulmonary clearance of NaF, with concurrent accumulation in bone and predominant urinary excretion within approximately 30 min after administration. These findings indicate that single airway exposure to NaF induces acute but reversible pulmonary inflammation despite rapid lung clearance, and call for evaluation under repeated or exposure-relevant conditions and in both sexes, as pulmonary toxicity was assessed only in male rats and only by intratracheal instillation.

Background and objectives: Occupational and environmental inhalation exposures, including high-aspect-ratio carbon nanotubes, can trigger pulmonary fibrosis (PF). The relationship between exposure-specific fibrogenic pathways (granulomatous inflammation versus diffuse epithelial injury) and lung microbiome dysbiosis remains incompletely understood. We therefore compared lung microbiome alterations in rat PF models induced by multi-walled carbon nanotubes (MWCNTs) and bleomycin. Materials and Methods: Female Wistar rats received a single intratracheal instillation of vehicle, MWCNTs (750 μg/rat), or bleomycin (1 mg/rat). At day 28, fibrosis and inflammation were evaluated by histopathology and bronchoalveolar lavage fluid (BALF) profiling. Lung microbial communities were characterized by 16S rRNA gene sequencing (V3–V4). Seventeen lung samples passed stringent quality control and were analyzed (control n = 5; bleomycin n = 7; MWCNT n = 5). Results: Both agents induced PF with increased profibrotic signaling, but with distinct pathological signatures: MWCNTs produced localized granulomatous lesions and a robust neutrophilic response (25% of BALF cells), whereas bleomycin caused diffuse interstitial remodeling. Bleomycin increased microbial richness (alpha diversity; p < 0.05) and significantly shifted community structure (beta diversity; p < 0.05), while MWCNT exposure showed comparatively limited changes in global diversity. The relative abundance of Pseudogracilibacillus (including P. marinus) was higher in the bleomycin group than in controls, whereas Facklamia tabacinasalis and Corynebacterium maris were more abundant in the MWCNT group. Across samples, Proteobacteria abundance was inversely correlated with BALF TGF-β, MCP-1, and neutrophil proportion. At the species level, Pseudogracilibacillus marinus was positively correlated with BALF TGF-β, while Facklamia tabacinasalis and Corynebacterium maris were positively correlated with MCP-1, CINC-3, and neutrophil proportion (Spearman; p < 0.05). Conclusions: Mechanistically distinct fibrogenic exposures generate exposure-linked lung microbiome signatures that track with host inflammatory and profibrotic responses. These signatures may support biomarker development for environmentally and occupationally relevant PF and motivate longitudinal and functional studies to clarify causality.