Pulmonary toxicity and translocation of gallium phosphide nanowires to secondary organs following pulmonary exposure in mice.

BACKGROUND: III-V semiconductor nanowires are envisioned as being integrated in optoelectronic devices in the near future. However, the perspective of mass production of these nanowires raises concern for human safety due to their asbestos- and carbon nanotube-like properties, including their high aspect ratio shape. Indeed, III-V nanowires have similar dimensions as Mitsui-7 multi-walled carbon nanotubes, which induce lung cancer by inhalation in rats. It is therefore urgent to investigate the toxicological effects following lung exposure to III-V nanowires prior to their use in industrial production, which entails risk of human exposure. Here, female C57BL/6J mice were exposed to 2, 6, and 18 µg (0.12, 0.35 and 1.1 mg/kg bw) of gallium phosphide (III-V) nanowires (99 nm diameter, 3.7 µm length) by intratracheal instillation and the toxicity was investigated 1, 3, 28 days and 3 months after exposure. Mitsui-7 multi-walled carbon nanotubes and carbon black Printex 90 nanoparticles were used as benchmark nanomaterials. RESULTS: Gallium phosphide nanowires induced genotoxicity in bronchoalveolar lavage cells and acute inflammation with eosinophilia observable both in bronchoalveolar lavage and lung tissue (1 and 3 days post-exposure). The inflammatory response was comparable to the response following exposure to Mitsui-7 multi-walled carbon nanotubes at similar dose levels. The nanowires underwent partial dissolution in the lung resulting in thinner nanowires, with an estimated in vivo half-life of 3 months. Despite the partial dissolution, nanowires were detected in lung, liver, spleen, kidney, uterus and brain 3 months after exposure. CONCLUSION: Pulmonary exposure to gallium phosphide nanowires caused similar toxicological effects as the multi-walled carbon nanotube Mitsui-7.

Protective effects of surfactant protein D treatment in 1,3-ß-glucan-modulated allergic inflammation.

Surfactant protein D (SP-D) is a pulmonary collectin important in lung immunity. SP-D-deficient mice (Sftpd(-/-)) are reported to be susceptible to ovalbumin (OVA)- and fungal allergen-induced pulmonary inflammation, while treatment with exogenous SP-D has therapeutic effects in such disease models. ß-Glucans are a diverse group of polysaccharides previously suggested to serve as fungal ligands for SP-D. We set out to investigate if SP-D could interact with 1,3-ß-glucan and attenuate allergic pulmonary inflammation in the presence of 1,3-ß-glucan. Allergic airway disease was induced in Sftpd(-/-) and Sftpd(+/+) mice by OVA sensitization and subsequent challenge with OVA, 1,3-ß-glucan, or OVA/1,3-ß-glucan together. Mice in the combined treatment group were further treated with a high dose of recombinant fragment of human SP-D (rfhSP-D). We demonstrated direct interaction between SP-D and 1,3-ß-glucan. OVA-induced mucous cell metaplasia was increased in Sftpd(-/-) mice, supporting previously reported protective effects of endogenous SP-D in allergy. OVA-induced parenchymal CCL11 levels and eosinophilic infiltration in bronchoalveolar lavage were unaffected by 1,3-ß-glucan, but were reversed with rfhSP-D treatment. 1,3-ß-Glucan treatment did, however, induce pulmonary neutrophilic infiltration and increased TNF-α levels in bronchoalveolar lavage, independently of OVA-induced allergy. This infiltration was also reversed by treatment with rfhSP-D. 1,3-ß-Glucan reduced OVA-induced mucous cell metaplasia, T helper 2 cytokines, and IFN-γ production. rfhSP-D treatment further reduced mucous metaplasia and T helper 2 cytokine secretion to background levels. In summary, rfhSP-D treatment resulted in attenuation of both allergic inflammation and 1,3-ß-glucan-mediated neutrophilic inflammation. Our data suggest that treatment with high-dose SP-D protects from mold-induced exacerbations of allergic asthma.

Addition of glycosylation to influenza A virus hemagglutinin modulates antibody-mediated recognition of H1N1 2009 pandemic viruses.

Seasonal influenza A viruses (IAV) originate from pandemic IAV and have undergone changes in antigenic structure, including addition of glycans to the viral hemagglutinin (HA). Glycans on the head of HA promote virus survival by shielding antigenic sites, but highly glycosylated seasonal IAV are inactivated by soluble lectins of the innate immune system. In 2009, human strains of pandemic H1N1 [A(H1N1)pdm] expressed a single glycosylation site (Asn(104)) on the head of HA. Since then, variants with additional glycosylation sites have been detected, and the location of these sites has been distinct to those of recent seasonal H1N1 strains. We have compared wild-type and reverse-engineered A(H1N1)pdm IAV with differing potential glycosylation sites on HA for sensitivity to collectins and to neutralizing Abs. Addition of a glycan (Asn(136)) to A(H1N1)pdm HA was associated with resistance to neutralizing Abs but did not increase sensitivity to collectins. Moreover, variants expressing Asn(136) showed enhanced growth in A(H1N1)pdm-vaccinated mice, consistent with evasion of Ab-mediated immunity in vivo. Thus, a fine balance exists regarding the optimal pattern of HA glycosylation to facilitate evasion of Ab-mediated immunity while maintaining resistance to lectin-mediated defenses of the innate immune system.

Pulmonary toxicity of synthetic amorphous silica - effects of porosity and copper oxide doping.

Materials can be modified for improved functionality. Our aim was to test whether pulmonary toxicity of silica nanomaterials is increased by the introduction of: a) porosity; and b) surface doping with CuO; and whether c) these modifications act synergistically. Mice were exposed by intratracheal instillation and for some doses also oropharyngeal aspiration to: 1) solid silica 100 nm; 2) porous silica 100 nm; 3) porous silica 100 nm with CuO doping; 4) solid silica 300 nm; 5) porous silica 300 nm; 6) solid silica 300 nm with CuO doping; 7) porous silica 300 nm with CuO doping; 8) CuO nanoparticles 9.8 nm; or 9) carbon black Printex 90 as benchmark. Based on a pilot study, dose levels were between 0.5 and 162 µg/mouse (0.2 and 8.1 mg/kg bw). Endpoints included pulmonary inflammation (neutrophil numbers in bronchoalveolar fluid), acute phase response, histopathology, and genotoxicity assessed by the comet assay, micronucleus test, and the gamma-H2AX assay. The porous silica materials induced greater pulmonary inflammation than their solid counterparts. A similar pattern was seen for acute phase response induction and histologic changes. This could be explained by a higher specific surface area per mass unit for the most toxic particles. CuO doping further increased the acute phase response normalized according to the deposited surface area. We identified no consistent evidence of synergism between surface area and CuO doping. In conclusion, porosity and CuO doping each increased the toxicity of silica nanomaterials and there was no indication of synergy when the modifications co-occurred.

Sub-chronic lung inflammation after airway exposures to Bacillus thuringiensis biopesticides in mice.

BACKGROUND: The aim of the present study was to assess possible health effects of airway exposures to Bacillus thuringiensis (Bt) based biopesticides in mice. Endpoints were lung inflammation evaluated by presence of inflammatory cells in bronchoalveolar lavage fluid (BALF), clearance of bacteria from the lung lumen and histological alterations of the lungs. Hazard identifications of the biopesticides were carried out using intratracheal (i.t.) instillation, followed by an inhalation study. The two commercial biopesticides used were based on the Bt. subspecies kurstaki and israelensis, respectively. Groups of BALB/c mice were i.t instilled with one bolus (3.5 × 105 or 3.4 × 106 colony forming units (CFU) per mouse) of either biopesticide. Control mice were instilled with sterile water. BALFs were collected and the inflammatory cells were counted and differentiated. The BALFs were also subjected to CFU counts. RESULTS: BALF cytology showed an acute inflammatory response dominated by neutrophils 24 hours after instillation of biopesticide. Four days after instillation, the neutrophil number was normalised and inflammation was dominated by lymphocytes and eosinophils, whereas 70 days after instillation, the inflammation was interstitially located with few inflammatory cells present in the lung lumen.Half of the instilled mice had remaining CFU recovered from BALF 70 days after exposure. To gain further knowledge with relevance for risk assessment, mice were exposed to aerosols of biopesticide one hour per day for 2 × 5 days. Each mouse received 1.9 × 104 CFU Bt israelensis or 2.3 × 103 CFU Bt kurstaki per exposure. Seventy days after end of the aerosol exposures, 3 out of 17 mice had interstitial lung inflammation. CFU could be recovered from 1 out of 10 mice 70 days after exposure to aerosolised Bt kurstaki. Plethysmography showed that inhalation of Bt aerosol did not induce airway irritation. CONCLUSIONS: Repeated low dose aerosol exposures to commercial Bt based biopesticides can induce sub-chronic lung inflammation in mice, which may be the first step in the development of chronic lung diseases. Inhalation of Bt aerosols does not induce airway irritation, which could explain why workers may be less inclined to use a filter mask during the application process, and are thereby less protected from exposure to Bt spores.

Pre-conceptional exposure to multiwalled carbon nanotubes suppresses antibody production in mouse offspring.

Prenatal particle exposure has been shown to increase allergic responses in offspring. Carbon nanotubes (CNTs) possess immunomodulatory properties, but it is unknown whether maternal exposure to CNTs interferes with offspring immune development. Here, C57Bl/6J female mice were intratracheally instilled with 67 of µg multiwalled CNTs on the day prior to mating. After weaning, tolerance and allergy responses were assessed in the offspring. Offspring of CNT-exposed (CNT offspring) and of sham-exposed dams (CTRL offspring) were intranasally exposed to ovalbumin (OVA) once weekly for 5 weeks to induce airway mucosal tolerance. Subsequent OVA sensitization and aerosol inhalation caused low or no OVA-specific IgE production and no inflammation. However, the CNT offspring presented with significantly lower OVA-specific IgG1 levels than CTRL offspring. In other groups of 5-week-old offspring, low-dose sensitization with OVA and subsequent OVA aerosol inhalation led to significantly lower OVA-specific IgG1 production in CNT compared to CTRL offspring. OVA-specific IgE and airway inflammation were non-significantly reduced in CNT offspring. The immunomodulatory effects of pre-gestational exposure to multiwalled CNTs were unexpected, but very consistent. The observations of suppressed antigen-specific IgG1 production may be of importance for infection or vaccination responses and warrant further investigation.

Alterations of the murine gut microbiome in allergic airway disease are independent of surfactant protein D.

BACKGROUND: SP-D is an important host defense lectin in innate immunity and SP-D deficient mice show several abnormal immune effects and are susceptible to allergen-induced airway disease. At the same time, host microbiome interactions play an important role in the development of allergic airway disease, and alterations to gut microbiota have been linked to airway disease through the gut-lung axis. Currently, it is unknown if the genotype (Sftpd-/- or Sftpd+/+) of the standard SP-D mouse model can affect the host microbiota to such an degree that it would overcome the cohousing effect on microbiota and interfere with the interpretation of immunological data from the model. Generally, little is known about the effect of the SP-D protein in itself and in combination with airway disease on the microbiota. In this study, we tested the hypothesis that microbiome composition would change with the lack of SP-D protein and presence of allergic airway disease in the widely used SP-D-deficient mouse model. RESULTS: We describe here for the first time the lung and gut microbiota of the SP-D mouse model with OVA induced allergic airway disease. After the challenge animals were killed and fecal samples were taken from the caecum and lungs were subjected to bronchoalveolar lavage for comparison of gut and lung microbiota by Illumina 16S rRNA gene sequencing. A significant community shift was observed in gut microbiota after challenge with OVA. However, the microbial communities were not significantly different between SP-D deficient and wild type mice from the same cages in either naïve or OVA treated animals. Wild type animals did however show the largest variation between mice. CONCLUSIONS: Our results show that the composition of the microbiota is not influenced by the SP-D deficient genotype under naïve or OVA induced airway disease. However, OVA sensitization and pulmonary challenge did alter the gut microbiota, supporting a bidirectional lung-gut crosstalk. Future mechanistic investigations of the influence of induced allergic airway disease on gut microbiota are warranted.

Cardiovascular health effects of oral and pulmonary exposure to multi-walled carbon nanotubes in ApoE-deficient mice.

Exposure to high aspect ratio nanomaterials, such as multi-walled carbon nanotubes (MWCNTs) may be associated with increased risk of atherosclerosis, pulmonary disease, and cancer. In the present study, we investigated the cardiovascular and pulmonary health effects of 10 weeks of repeated oral or pulmonary exposures to MWCNTs (4 or 40µg each week) in Apolipoprotein E-deficient (ApoE-/-) mice fed a Western-type diet. Intratracheal instillation of MWCNTs was associated with oxidative damage to DNA in lung tissue and elevated levels of lipid peroxidation products in plasma, whereas the exposure only caused a modest pulmonary inflammation in terms of increased numbers of lymphocytes in bronchoalveolar lavage fluid. Ultrasound imaging in live animals revealed an increase in the inner and outer wall thickness of the aortic arch at 10 weeks after pulmonary exposure to MWCNTs, which may suggest artery remodelling. However, we did not find accelerated plaque progression in the aorta or the brachiocephalic artery by histopathology. Furthermore, repeated oral exposure to MWCNTs did not cause changes in the composition of gut microbiota of exposed mice. Collectively, this study indicates that repeated pulmonary exposure to MWCNTs was associated with oxidative stress, whereas cardiovascular effects encompassed remodelling of the aorta wall.