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Formation of reactive oxygen species in rat epithelial cells upon stimulation with fly ash

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Abstract

Fly ash was used as a model for ambient particulate matter which is under suspicion to cause adverse pulmonary health effects. The fly ash was pre-sized and contained only particles < 20 μm including an ultrafine fraction (< 100 nm) that contributed 31% to the particle number. In our study, we investigated the influence of fly ash on the promotion of early inflammatory reactions like the formation of reactive oxygen species (ROS) in rat lung epithelial cells (RLE-6TN). Furthermore, we determined the formation of nitric oxide (NO). The cells show a clear dose-response relationship concerning the formation of ROS with regard to the mass of particles applied. Lipopolysaccharide (LPS) added as a co-stimulus did not increase the formation of ROS induced by fly ash. Furthermore, in LPS (01 μg/ml) and tumour necrosis factor-alpha (TNF-alpha; 1 ng/ml) pre-treated cells no increase in reactive oxygen species comparable to fly ash alone is observable. In presence of the metal chelator, desferrioxamine (DFO), ROS formation can be significantly reduced. Neither fly ash nor LPS induced a significant NO release in RLE-6TN cells.

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Abbreviations

BSA:

Bovine serum albumin

DCF:

2′-7′-dichlorofluorescein

DFO:

Desferrioxamine mesylate

DMEM:

Dulbecco’s Modified Eagle Medium

FCS:

fetal calf serum

HBSS:

Hank’s balanced salt solution

H2DCF:

2′-7′-dichlorodihydro-fluorescein

IFN-gamma:

interferon-gamma

iNOS:

inducible nitric oxide synthase

MAF98:

fly ash used in this study

LBP:

lipopolysaccharide binding protein

LPS:

lipopolysaccharide

MIP-2:

macrophage inflammatory protein-2

NO:

nitric oxide

ROS:

reactive oxygen species

TNF-alpha:

tumour necrosis factor-alpha

TPA:

12-O-tetradecanoylphorbol-13-acetate

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Authors and Affiliations

  1. Institute of Toxicology and Genetics, Forschungszentrum Karlsruhe, D-76021, Karlsruhe, Germany

    K. Voelkel, H. F. Krug & S. Diabaté

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  1. K. Voelkel
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  2. H. F. Krug
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  3. S. Diabaté
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Correspondence to K. Voelkel.

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Voelkel, K., Krug, H.F. & Diabaté, S. Formation of reactive oxygen species in rat epithelial cells upon stimulation with fly ash. J. Biosci. 28, 51–55 (2003). https://doi.org/10.1007/BF02970131

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