Limits...
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.

Show MeSH

Related in: MedlinePlus

ZnO deposition onto cell cultures, out of differently concentrated suspensions, measured by atomic absorption spectroscopy. The deposition reaches a plateau over the first 4 h of incubation. 5 – 60 ppm suspensions correspond to an aerosol – exposure for 22 – 90 seconds. Deposition efficiency increases with concentration. Data are presented as mean ± SD.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3585858&req=5

Figure 7: ZnO deposition onto cell cultures, out of differently concentrated suspensions, measured by atomic absorption spectroscopy. The deposition reaches a plateau over the first 4 h of incubation. 5 – 60 ppm suspensions correspond to an aerosol – exposure for 22 – 90 seconds. Deposition efficiency increases with concentration. Data are presented as mean ± SD.

Mentions: Equivalent particle doses are a prerequisite for a comparison of aerosol – and suspension exposure scenarios. The depletion of zinc in the cellular supernatant, ranging from 5–60 ppm (parts per million, = μg/mL culture medium), was assessed over time (Additional file1: Figure S2), and the obtained data were expressed as mass deposition per culture surface (Figure7). A plateau was reached over the first two hours, with a mean deposited mass of 0.5 (SD 0.4, n = 4), 2.2 (SD 0.9, n = 4), 5.8 (SD 1.6, n = 4) and 13.6 (SD 2.1, n =4) μg/cm2 for 5, 15, 30 and 60 ppm suspensions after 4 h exposure. This is 37.6, 55.8, 73.8 and 86.0% of the maximal possible deposition, which was calculated to 1.3, 3.9, 7.9 and 15.8 μg/cm2. A dose range of 5 – 30 ppm was found to be comparable, following 22 – 90 sec particle production (1.3 – 6.1 μg/cm2) in the aerosol scenario. Basing on this knowledge, suspension experiments were performed with ZnO suspensions of 0 – 30 ppm, as previously investigated by Brunner et al.[13], with additional 60 and 80 ppm suspensions as “high dose” controls.


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)

ZnO deposition onto cell cultures, out of differently concentrated suspensions, measured by atomic absorption spectroscopy. The deposition reaches a plateau over the first 4 h of incubation. 5 – 60 ppm suspensions correspond to an aerosol – exposure for 22 – 90 seconds. Deposition efficiency increases with concentration. Data are presented as mean ± SD.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 7: ZnO deposition onto cell cultures, out of differently concentrated suspensions, measured by atomic absorption spectroscopy. The deposition reaches a plateau over the first 4 h of incubation. 5 – 60 ppm suspensions correspond to an aerosol – exposure for 22 – 90 seconds. Deposition efficiency increases with concentration. Data are presented as mean ± SD.
Mentions: Equivalent particle doses are a prerequisite for a comparison of aerosol – and suspension exposure scenarios. The depletion of zinc in the cellular supernatant, ranging from 5–60 ppm (parts per million, = μg/mL culture medium), was assessed over time (Additional file1: Figure S2), and the obtained data were expressed as mass deposition per culture surface (Figure7). A plateau was reached over the first two hours, with a mean deposited mass of 0.5 (SD 0.4, n = 4), 2.2 (SD 0.9, n = 4), 5.8 (SD 1.6, n = 4) and 13.6 (SD 2.1, n =4) μg/cm2 for 5, 15, 30 and 60 ppm suspensions after 4 h exposure. This is 37.6, 55.8, 73.8 and 86.0% of the maximal possible deposition, which was calculated to 1.3, 3.9, 7.9 and 15.8 μg/cm2. A dose range of 5 – 30 ppm was found to be comparable, following 22 – 90 sec particle production (1.3 – 6.1 μg/cm2) in the aerosol scenario. Basing on this knowledge, suspension experiments were performed with ZnO suspensions of 0 – 30 ppm, as previously investigated by Brunner et al.[13], with additional 60 and 80 ppm suspensions as “high dose” controls.

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