Toxicity evaluation following pulmonary exposure to an as-manufactured dispersed boron nitride nanotube (BNNT) material in vivo

Abstract Boron nitride nanotubes (BNNT) are multi-walled nanotubes composed of hexagonal B N bonds and possess many unique physical and chemical properties, creating a rapidly expanding market for this newly emerging nanomaterial which is still primarily in the research and development stage. The shape and high aspect ratio give rise to concern for the potential toxicity that may be associated with pulmonary exposure, especially in an occupational setting. The goal of this study was to assess lung toxicity using an in vivo time course model. The sample was manufactured to be 5 nm wide and up to 200 μm long, with ~50% purity covalently bound with hexagonal boron nitride (hBN) in the sample. Following preparation for in vivo studies, sonication of the material disrupted the longer tubes in the complex and the size distribution in dispersion medium (DM) of the structures was 13–23 nm in diameter and 0.6–1.6 μm in length. Male C57BL/6 J mice were exposed to 4 or 40 μg of BNNT or DM (vehicle control) by a single oropharyngeal aspiration. Pulmonary and systemic toxicity were investigated at 4 h, 1 d, 7 d, 1 mo and 2 mo post-exposure. Bronchoalveolar lavage (BAL) studies determined pulmonary inflammation (neutrophil influx) and cytotoxicity (lactate dehydrogenase activity) occurred at early time points and peaked at 7 d post-exposure in the high dose group. Histopathological analysis showed a minimal level of inflammatory cell infiltration in the high dose group with resolution over time and no fibrosis, and lung clearance analysis showed ~50% of the material cleared over the time course. The expression of inflammatory- and acute phase response-associated genes in the lung and liver were significantly increased by the high dose at 4 h and 1 d post-exposure. The increases in lung gene expression of Cxcl2, Ccl2, Il6, Ccl22, Ccl11, and Spp1 were significant up to 2 mo but decreased with time. The low dose exposure did not result in significant changes in any toxicological parameters measured. In summary, the BNNT-hBN sample used in this study caused acute pulmonary inflammation and injury at the higher dose, which peaked by 7 d post-exposure and showed resolution over time. Further studies are needed to determine if physicochemical properties and purity will impact the toxicity profile of BNNT and to investigate the underlying mechanisms of BNNT toxicity.