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Differential proteomic analysis of mouse macrophages exposed to adsorbate-loaded heavy fuel oil derived combustion particles using an automated sample-preparation workflow

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Abstract

Ship diesel combustion particles are known to cause broad cytotoxic effects and thereby strongly impact human health. Particles from heavy fuel oil (HFO) operated ships are considered as particularly dangerous. However, little is known about the relevant components of the ship emission particles. In particular, it is interesting to know if the particle cores, consisting of soot and metal oxides, or the adsorbate layers, consisting of semi- and low-volatile organic compounds and salts, are more relevant. We therefore sought to relate the adsorbates and the core composition of HFO combustion particles to the early cellular responses, allowing for the development of measures that counteract their detrimental effects. Hence, the semi-volatile coating of HFO-operated ship diesel engine particles was removed by stepwise thermal stripping using different temperatures. RAW 264.7 macrophages were exposed to native and thermally stripped particles in submersed culture. Proteomic changes were monitored by two different quantitative mass spectrometry approaches, stable isotope labeling by amino acids in cell culture (SILAC) and dimethyl labeling. Our data revealed that cells reacted differently to native or stripped HFO combustion particles. Cells exposed to thermally stripped particles showed a very differential reaction with respect to the composition of the individual chemical load of the particle. The cellular reactions of the HFO particles included reaction to oxidative stress, reorganization of the cytoskeleton and changes in endocytosis. Cells exposed to the 280 °C treated particles showed an induction of RNA-related processes, a number of mitochondria-associated processes as well as DNA damage response, while the exposure to 580 °C treated HFO particles mainly induced the regulation of intracellular transport. In summary, our analysis based on a highly reproducible automated proteomic sample-preparation procedure shows a diverse cellular response, depending on the soot particle composition. In particular, it was shown that both the molecules of the adsorbate layer as well as particle cores induced strong but different effects in the exposed cells.

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Abbreviations

DML:

Dimethyl labeling

GC:

Gas chromatography

GO:

Gene ontology

HFO:

Heavy fuel oil

HFO280:

Heavy fuel oil, 280 °C treatment

HFO580:

Heavy fuel oil, 580 °C treatment

LOD:

Limit of detection

PAH:

Polycyclic aromatic hydrocarbons

PM:

Particulate matter

SILAC:

Stable isotope labeling by amino acids in cell culture

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Acknowledgments

This work was supported by funds of the Helmholtz virtual institute for complex Molecular Systems in Environmental Health (HICE). We thank Hanns Paur from the KIT, Karlsruhe, Germany for the loan of the CAROLA electrostatic precipitator and Patrick Beaudette as well as Daniel Perez-Hernandez for carefully reading the paper and fruitful discussions. We thank CTC and the Axel Semrau GmbH for the close collaboration in adapting the automation procedures for the CTC-PAL/Chronos setup.

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The authors declare no conflict of interest.

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Correspondence to Gunnar Dittmar.

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Published in the topical collection Aerosols and Health with guest editor Ralf Zimmermann.

Tamara Kanashova and Oliver Popp contributed equally to this work.

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Kanashova, T., Popp, O., Orasche, J. et al. Differential proteomic analysis of mouse macrophages exposed to adsorbate-loaded heavy fuel oil derived combustion particles using an automated sample-preparation workflow. Anal Bioanal Chem 407, 5965–5976 (2015). https://doi.org/10.1007/s00216-015-8595-4

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  • DOI: https://doi.org/10.1007/s00216-015-8595-4

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