Procedural application of mode-of-action and human relevance analysis: styrene-induced lung tumors in mice.

Risk assessment of human health hazards has traditionally relied on experiments that use animal models. Although exposure studies in rats and mice are a major basis for determining risk in many cases, observations made in animals do not always reflect health hazards in humans due to differences in biology. In this critical review, we use the mode-of-action (MOA) human relevance framework to assess the likelihood that bronchiolar lung tumors observed in mice chronically exposed to styrene represent a plausible tumor risk in humans. Using available datasets, we analyze the weight-of-evidence 1) that styrene-induced tumors in mice occur through a MOA based on metabolism of styrene by Cyp2F2; and 2) whether the hypothesized key event relationships are likely to occur in humans. This assessment describes how the five modified Hill causality considerations support that a Cyp2F2-dependent MOA causing lung tumors is active in mice, but only results in tumorigenicity in susceptible strains. Comparison of the key event relationships assessed in the mouse was compared to an analogous MOA hypothesis staged in the human lung. While some biological concordance was recognized between key events in mice and humans, the MOA as hypothesized in the mouse appears unlikely in humans due to quantitative differences in the metabolic capacity of the airways and qualitative uncertainties in the toxicological and prognostic concordance of pre-neoplastic and neoplastic lesions arising in either species. This analysis serves as a rigorous demonstration of the framework's utility in increasing transparency and consistency in evidence-based assessment of MOA hypotheses in toxicological models and determining relevance to human health.

Genetic susceptibility to toxicologic lung responses among inbred mouse strains following exposure to carbon nanotubes and profiling of underlying gene networks.

The risk of human exposure to fiber nanoparticles has risen in recent years due to increases in the manufacture and utilization of carbon nanotubes (CNTs). CNTs are present as airborne particulates in occupational settings and their hazard potential has been demonstrated in experimental lung exposure studies using inbred mouse strains. However, it is not known whether different inbred strains differ in lung responses to CNTs by virtue of their genetics. In this work, common inbred strains (BALB/c, C57Bl/6, DBA/2, and C3H/He) were exposed to CNTs via oropharyngeal aspiration and lung histology and bronchoalveolar lavage (BAL) samples were evaluated over 28days with the objective of evaluating sensitivity/resistance among strains. C57Bl/6 mice developed significantly more extensive type II pneumocyte (T2P) hyperplasia and alveolar infiltrate compared to DBA/2 mice, which were resistant. Surprisingly, DBA/2 but not C57Bl/6 mice were extremely sensitive to increases in leukocytes recovered in BAL fluid. Underlying global gene expression patterns in the two strains were compared using mRNA sequencing to investigate regulatory networks associated with the different effects. The impact of exposure on gene networks regulating various aspects of immune response and cell survival was limited in DBA/2 mice compared to C57Bl/6. Investigation of B6D2F1 (C57Bl/6×DBA/2 hybrid) mice demonstrated inheritance of sensitivity to CNT exposures in regard to toxicologic lung pathology and BAL leukocyte accumulations. These findings demonstrate a genetic basis of susceptibility to CNT particle exposures and both inform the use of inbred mouse models and suggest the likelihood of differences in genetic susceptibility among humans.

Carbon Nanotube and Asbestos Exposures Induce Overlapping but Distinct Profiles of Lung Pathology in Non-Swiss Albino CF-1 Mice.

Carbon nanotubes (CNTs) are emerging as important occupational and environmental toxicants owing to their increasing prevalence and potential to be inhaled as airborne particles. CNTs are a concern because of their similarities to asbestos, which include fibrous morphology, high aspect ratio, and biopersistence. Limitations in research models have made it difficult to experimentally ascertain the risk of CNT exposures to humans and whether these may lead to lung diseases classically associated with asbestos, such as mesothelioma and fibrosis. In this study, we sought to comprehensively compare profiles of lung pathology in mice following repeated exposures to multiwall CNTs or crocidolite asbestos (CA). We show that both exposures resulted in granulomatous inflammation and increased interstitial collagen; CA exposures caused predominantly bronchoalveolar hyperplasia, whereas CNT exposures caused alveolar hyperplasia of type II pneumocytes (T2Ps). T2Ps isolated from CNT-exposed lungs were found to have upregulated proinflammatory genes, including interleukin 1ß (IL-1ß), in contrast to those from CA exposed. Immunostaining in tissue showed that while both toxicants increased IL-1ß protein expression in lung cells, T2P-specific IL-1ß increases were greater following CNT exposure. These results suggest related but distinct mechanisms of action by CNTs versus asbestos which may lead to different outcomes in the 2 exposure types.

MyD88 mediates in vivo effector functions of alveolar macrophages in acute lung inflammatory responses to carbon nanotube exposure.

Carbon nanotubes (CNTs) are rapidly emerging as high-priority occupational toxicants. CNT powders contain fibrous particles that aerosolize readily in places of manufacture and handling, posing an inhalation risk for workers. Studies using animal models indicate that lung exposure to CNTs causes prolonged inflammatory responses and diffuse alveolar injury. The mechanisms governing CNT-induced lung inflammation are not fully understood but have been suggested to involve alveolar macrophages (AMs). In the current study, we sought to systematically assess the effector role of AMs in vivo in the induction of lung inflammatory responses to CNT exposures and investigate their cell type-specific mechanisms. Multi-wall CNTs characterized for various physicochemical attributes were used as the CNT type. Using an AM-specific depletion and repopulation approach in a mouse model, we unambiguously demonstrated that AMs are major effector cells necessary for the in vivo elaboration of CNT-induced lung inflammation. We further investigated in vitro AM responses and identified molecular targets which proved critical to pro-inflammatory responses in this model, namely MyD88 as well as MAPKs and Ca(2+)/CamKII. We further demonstrated that MyD88 inhibition in donor AMs abrogated their capacity to reconstitute CNT-induced inflammation when adoptively transferred into AM-depleted mice. Taken together, this is the first in vivo demonstration that AMs act as critical effector cell types in CNT-induced lung inflammation and that MyD88 is required for this in vivo effector function. AMs and their cell type-specific mechanisms may therefore represent potential targets for future therapeutic intervention of CNT-related lung injury.