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Effects of flame made zinc oxide particles in human lung cells - a comparison of aerosol and suspension exposures.

Raemy DO, Grass RN, Stark WJ, Schumacher CM, Clift MJ, Gehr P, Rothen-Rutishauser B - Part Fibre Toxicol (2012)

Bottom Line: Such an approach however, does not reflect particle inhalation.No direct effects could be attributed to ZnO particles.Exposure to suspensions represents a valuable complementary method and allows investigations on particle-associated toxicity by excluding all gas-derived effects.

View Article: PubMed Central - HTML - PubMed

Affiliation: Adolphe Merkle Institute, Bionanomaterials, University of Fribourg, Rte de l'Ancienne Papeterie, P.O. Box 209, CH-1732, Marly, Switzerland.

ABSTRACT

Background: Predominantly, studies of nanoparticle (NPs) toxicology in vitro are based upon the exposure of submerged cell cultures to particle suspensions. Such an approach however, does not reflect particle inhalation. As a more realistic simulation of such a scenario, efforts were made towards direct delivery of aerosols to air-liquid-interface cultivated cell cultures by the use of aerosol exposure systems.This study aims to provide a direct comparison of the effects of zinc oxide (ZnO) NPs when delivered as either an aerosol, or in suspension to a triple cell co-culture model of the epithelial airway barrier. To ensure dose-equivalence, ZnO-deposition was determined in each exposure scenario by atomic absorption spectroscopy. Biological endpoints being investigated after 4 or 24h incubation include cytotoxicity, total reduced glutathione, induction of antioxidative genes such as heme-oxygenase 1 (HO-1) as well as the release of the (pro)-inflammatory cytokine TNFα.

Results: Off-gases released as by-product of flame ZnO synthesis caused a significant decrease of total reduced GSH and induced further the release of the cytokine TNFα, demonstrating the influence of the gas phase on aerosol toxicology. No direct effects could be attributed to ZnO particles. By performing suspension exposure to avoid the factor "flame-gases", particle specific effects become apparent. Other parameters such as LDH and HO-1 were not influenced by gaseous compounds: Following aerosol exposure, LDH levels appeared elevated at both timepoints and the HO-1 transcript correlated positively with deposited ZnO-dose. Under submerged conditions, the HO-1 induction scheme deviated for 4 and 24h and increased extracellular LDH was found following 24h exposure.

Conclusion: In the current study, aerosol and suspension-exposure has been compared by exposing cell cultures to equivalent amounts of ZnO. Both exposure strategies differ fundamentally in their dose-response pattern. Additional differences can be found for the factor time: In the aerosol scenario, parameters tend to their maximum already after 4h of exposure, whereas under submerged conditions, effects appear most pronounced mainly after 24h. Aerosol exposure provides information about the synergistic interplay of gaseous and particulate phase of an aerosol in the context of inhalation toxicology. Exposure to suspensions represents a valuable complementary method and allows investigations on particle-associated toxicity by excluding all gas-derived effects.

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Related in: MedlinePlus

Real-time PCR on SOD1 and HO-1 in suspension exposed cultures. No regulation of SOD1 was found. In contrast, HO-1 is constantly induced (over the entire dose range) after 4 h incubation. Furthermore, a dose dependent increase was observed after 24 h. White, open bars represent mean values after 4 h post – incubation, black bars indicate 24 h timepoints. Individual values are expressed as data points.
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Figure 10: Real-time PCR on SOD1 and HO-1 in suspension exposed cultures. No regulation of SOD1 was found. In contrast, HO-1 is constantly induced (over the entire dose range) after 4 h incubation. Furthermore, a dose dependent increase was observed after 24 h. White, open bars represent mean values after 4 h post – incubation, black bars indicate 24 h timepoints. Individual values are expressed as data points.

Mentions: SOD1 remained on basal level for all conditions and timepoints (Figure10). After 4 h exposure, HO-1 values were 1.0 (SD 0.0), 4.2 (SD 2.3), 4.4 (SD 2.0), 5.0 (SD 1.7), 5.0 (SD 1.3) and 4.6 (SD 1.5) for concentrations of 0 ppm to 80 ppm (all n = 4). No significant effect was found by the Friedman test (p>0.05). The corresponding data for 24 h were 1.0 (SD 0.0), 0.8 (SD 0.2), 1.6 (SD 0.6), 3.6 (SD 0.6), 6.5 (SD 2.4) and 6.2 (SD 1.7) (all n = 4).). After 24 h exposure, a significant induction was demonstrated by the Friedman test (p = 0.012) and a significant dose dependent trend and correlation of dose and HO-1 transcript was found by the Jonckheere – Terpstra – and Kendall’s tau analysis (p = 0.000, τ = 0.776). The LPS positive control was determined as 1.8 (SD 1.5) and 6.1 (SD 0.8) for 4 and 24 h (n = 2).


Effects of flame made zinc oxide particles in human lung cells - a comparison of aerosol and suspension exposures.

Raemy DO, Grass RN, Stark WJ, Schumacher CM, Clift MJ, Gehr P, Rothen-Rutishauser B - Part Fibre Toxicol (2012)

Real-time PCR on SOD1 and HO-1 in suspension exposed cultures. No regulation of SOD1 was found. In contrast, HO-1 is constantly induced (over the entire dose range) after 4 h incubation. Furthermore, a dose dependent increase was observed after 24 h. White, open bars represent mean values after 4 h post – incubation, black bars indicate 24 h timepoints. Individual values are expressed as data points.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3585858&req=5

Figure 10: Real-time PCR on SOD1 and HO-1 in suspension exposed cultures. No regulation of SOD1 was found. In contrast, HO-1 is constantly induced (over the entire dose range) after 4 h incubation. Furthermore, a dose dependent increase was observed after 24 h. White, open bars represent mean values after 4 h post – incubation, black bars indicate 24 h timepoints. Individual values are expressed as data points.
Mentions: SOD1 remained on basal level for all conditions and timepoints (Figure10). After 4 h exposure, HO-1 values were 1.0 (SD 0.0), 4.2 (SD 2.3), 4.4 (SD 2.0), 5.0 (SD 1.7), 5.0 (SD 1.3) and 4.6 (SD 1.5) for concentrations of 0 ppm to 80 ppm (all n = 4). No significant effect was found by the Friedman test (p>0.05). The corresponding data for 24 h were 1.0 (SD 0.0), 0.8 (SD 0.2), 1.6 (SD 0.6), 3.6 (SD 0.6), 6.5 (SD 2.4) and 6.2 (SD 1.7) (all n = 4).). After 24 h exposure, a significant induction was demonstrated by the Friedman test (p = 0.012) and a significant dose dependent trend and correlation of dose and HO-1 transcript was found by the Jonckheere – Terpstra – and Kendall’s tau analysis (p = 0.000, τ = 0.776). The LPS positive control was determined as 1.8 (SD 1.5) and 6.1 (SD 0.8) for 4 and 24 h (n = 2).

Bottom Line: Such an approach however, does not reflect particle inhalation.No direct effects could be attributed to ZnO particles.Exposure to suspensions represents a valuable complementary method and allows investigations on particle-associated toxicity by excluding all gas-derived effects.

View Article: PubMed Central - HTML - PubMed

Affiliation: Adolphe Merkle Institute, Bionanomaterials, University of Fribourg, Rte de l'Ancienne Papeterie, P.O. Box 209, CH-1732, Marly, Switzerland.

ABSTRACT

Background: Predominantly, studies of nanoparticle (NPs) toxicology in vitro are based upon the exposure of submerged cell cultures to particle suspensions. Such an approach however, does not reflect particle inhalation. As a more realistic simulation of such a scenario, efforts were made towards direct delivery of aerosols to air-liquid-interface cultivated cell cultures by the use of aerosol exposure systems.This study aims to provide a direct comparison of the effects of zinc oxide (ZnO) NPs when delivered as either an aerosol, or in suspension to a triple cell co-culture model of the epithelial airway barrier. To ensure dose-equivalence, ZnO-deposition was determined in each exposure scenario by atomic absorption spectroscopy. Biological endpoints being investigated after 4 or 24h incubation include cytotoxicity, total reduced glutathione, induction of antioxidative genes such as heme-oxygenase 1 (HO-1) as well as the release of the (pro)-inflammatory cytokine TNFα.

Results: Off-gases released as by-product of flame ZnO synthesis caused a significant decrease of total reduced GSH and induced further the release of the cytokine TNFα, demonstrating the influence of the gas phase on aerosol toxicology. No direct effects could be attributed to ZnO particles. By performing suspension exposure to avoid the factor "flame-gases", particle specific effects become apparent. Other parameters such as LDH and HO-1 were not influenced by gaseous compounds: Following aerosol exposure, LDH levels appeared elevated at both timepoints and the HO-1 transcript correlated positively with deposited ZnO-dose. Under submerged conditions, the HO-1 induction scheme deviated for 4 and 24h and increased extracellular LDH was found following 24h exposure.

Conclusion: In the current study, aerosol and suspension-exposure has been compared by exposing cell cultures to equivalent amounts of ZnO. Both exposure strategies differ fundamentally in their dose-response pattern. Additional differences can be found for the factor time: In the aerosol scenario, parameters tend to their maximum already after 4h of exposure, whereas under submerged conditions, effects appear most pronounced mainly after 24h. Aerosol exposure provides information about the synergistic interplay of gaseous and particulate phase of an aerosol in the context of inhalation toxicology. Exposure to suspensions represents a valuable complementary method and allows investigations on particle-associated toxicity by excluding all gas-derived effects.

Show MeSH
Related in: MedlinePlus