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Toxic Effects of the Major Components of Diesel Exhaust in Human Alveolar Basal Epithelial Cells (A549)

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ABSTRACT

We investigated the toxicity of benzo[a]pyrene (B[a]P), 1-nitropyrene (1-NP) and 3-nitrobenzanthrone (3-NBA) in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 μM), 1-NP (1 and 10 μM) and 3-NBA (0.5 and 5 μM). Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH) nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties.

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Levels of protein carbonyls relative to the control (%) induced after treatment of A549 cells with benzo[a]pyrene, 1-nitropyrene and 3-nitrobenzanthrone for 4 h (A) and 24 h (B). Data represent mean ± standard deviation from two triplicates from two independent experiments (analyzed as n = 6). Asterisks denote significant (* p < 0.05) changes. The control level of protein carbonyls is represented by an emboldened horizontal line. 1-nitropyrene (10 µM, 4 h; 1 µM and 10 µM, 24 h) significantly increased protein carbonyl levels.
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ijms-17-01393-f005: Levels of protein carbonyls relative to the control (%) induced after treatment of A549 cells with benzo[a]pyrene, 1-nitropyrene and 3-nitrobenzanthrone for 4 h (A) and 24 h (B). Data represent mean ± standard deviation from two triplicates from two independent experiments (analyzed as n = 6). Asterisks denote significant (* p < 0.05) changes. The control level of protein carbonyls is represented by an emboldened horizontal line. 1-nitropyrene (10 µM, 4 h; 1 µM and 10 µM, 24 h) significantly increased protein carbonyl levels.

Mentions: Analyses of oxidative damage (Figure 3, Figure 4 and Figure 5) indicate relatively weak and non-consistent effects of test compounds on markers of oxidative damage. None of the time intervals and test (non-cytotoxic) doses of B[a]P increased levels of oxidative damage to DNA (8-oxodG, Figure 3A,B), lipids (15-F2t-IsoP, Figure 4A,B) or proteins (protein carbonyls, Figure 5A,B) significantly. Most of the significant biological effects could be ascribed to 1-NP treatment. This compound increased levels of lipid peroxidation after 4-h treatment (Figure 4A) and induced protein oxidation after 4-h and 24-h incubation (Figure 5A,B). However, no significant effect on DNA oxidation (Figure 3A,B) or lipid peroxidation after 24-h treatment (Figure 4B) was detected. 3-NBA significantly induced lipid peroxidation after 24-h treatment; levels of 15-F2t-IsoP increased to 200% of the relative control level, and were comparable for both test concentrations (Figure 4B). Levels of lipid peroxidation after 4-h treatment with 3-NBA were elevated, but the increase was not significant (Figure 4A). We did not detect an effect of this compound on oxidation of DNA or protein (Figure 3A,B and Figure 5A,B).


Toxic Effects of the Major Components of Diesel Exhaust in Human Alveolar Basal Epithelial Cells (A549)
Levels of protein carbonyls relative to the control (%) induced after treatment of A549 cells with benzo[a]pyrene, 1-nitropyrene and 3-nitrobenzanthrone for 4 h (A) and 24 h (B). Data represent mean ± standard deviation from two triplicates from two independent experiments (analyzed as n = 6). Asterisks denote significant (* p < 0.05) changes. The control level of protein carbonyls is represented by an emboldened horizontal line. 1-nitropyrene (10 µM, 4 h; 1 µM and 10 µM, 24 h) significantly increased protein carbonyl levels.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5037673&req=5

ijms-17-01393-f005: Levels of protein carbonyls relative to the control (%) induced after treatment of A549 cells with benzo[a]pyrene, 1-nitropyrene and 3-nitrobenzanthrone for 4 h (A) and 24 h (B). Data represent mean ± standard deviation from two triplicates from two independent experiments (analyzed as n = 6). Asterisks denote significant (* p < 0.05) changes. The control level of protein carbonyls is represented by an emboldened horizontal line. 1-nitropyrene (10 µM, 4 h; 1 µM and 10 µM, 24 h) significantly increased protein carbonyl levels.
Mentions: Analyses of oxidative damage (Figure 3, Figure 4 and Figure 5) indicate relatively weak and non-consistent effects of test compounds on markers of oxidative damage. None of the time intervals and test (non-cytotoxic) doses of B[a]P increased levels of oxidative damage to DNA (8-oxodG, Figure 3A,B), lipids (15-F2t-IsoP, Figure 4A,B) or proteins (protein carbonyls, Figure 5A,B) significantly. Most of the significant biological effects could be ascribed to 1-NP treatment. This compound increased levels of lipid peroxidation after 4-h treatment (Figure 4A) and induced protein oxidation after 4-h and 24-h incubation (Figure 5A,B). However, no significant effect on DNA oxidation (Figure 3A,B) or lipid peroxidation after 24-h treatment (Figure 4B) was detected. 3-NBA significantly induced lipid peroxidation after 24-h treatment; levels of 15-F2t-IsoP increased to 200% of the relative control level, and were comparable for both test concentrations (Figure 4B). Levels of lipid peroxidation after 4-h treatment with 3-NBA were elevated, but the increase was not significant (Figure 4A). We did not detect an effect of this compound on oxidation of DNA or protein (Figure 3A,B and Figure 5A,B).

View Article: PubMed Central - PubMed

ABSTRACT

We investigated the toxicity of benzo[a]pyrene (B[a]P), 1-nitropyrene (1-NP) and 3-nitrobenzanthrone (3-NBA) in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 &mu;M), 1-NP (1 and 10 &mu;M) and 3-NBA (0.5 and 5 &mu;M). Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH) nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties.

No MeSH data available.