Oxidative DNA damage induced by nitrotyrosine, a biomarker of inflammation

doi:10.1016/j.bbrc.2004.02.022

Biochemical and Biophysical Research Communications

Volume 316, Issue 1, 26 March 2004, Pages 123-128

https://doi.org/10.1016/j.bbrc.2004.02.022 Get rights and content

Abstract

Inflammation has been postulated as a risk factor for several cancers. 3-Nitrotyrosine is a biochemical marker for inflammation. We investigated the ability of nitrotyrosine and nitrotyrosine-containing peptides (nitroY-peptide) to induce DNA damage by the experiments using ³²P-labeled DNA fragments obtained from the human p53 tumor suppressor gene and an HPLC-electrochemical detector. Nitrotyrosine and nitroY-peptide caused Cu(II)-dependent DNA damage in the presence of P450 reductase, which is considered to yield nitroreduction. Catalase inhibited DNA damage, suggesting the involvement of H₂O₂. Nitrotyrosine and nitroY-peptide increased 8-oxo-7,8-dihydro-2^′-deoxyguanosine (8-oxodG) formation, an indicator of oxidative DNA damage. Nitrotyrosine-containing peptides of histone induced 8-oxodG formation more efficiently than free nitrotyrosine. We propose the possibility that nitrotyrosine-induced H₂O₂ formation and DNA damage contribute to inflammation-associated carcinogenesis.

Section snippets

Materials and methods

Materials. Restriction enzymes (EcoRI, MroI, and ApaI) and calf intestine phosphatase were purchased from Boehringer–Mannheim (Germany). Restriction enzymes (HindIII and AvaI) and T₄ polynucleotide kinase were purchased from New England Biolabs. [γ-³²P]ATP (222 TBq/mmol) was from New England Nuclear. Superoxide dismutase (SOD, 3000 U/mg from bovine erythrocytes) and catalase (45,000 U/mg from bovine liver) were from Sigma Chemical. Nitrotyrosine-containing peptides (nitroY-peptide) were supplied

Damage to ³²P-labeled DNA fragments by nitrotyrosine in the presence of P450 reductase, NADPH, and Cu(II)

Free nitrotyrosine and nitrotyrosine-peptides of histone caused Cu(II)-mediated DNA damage when they were treated with P450 reductase (Fig. 1A). Without P450 reductase, free nitrotyrosine and nitroY-peptides caused no DNA damage even in the presence of Cu(II) (data not shown). In the absence of Cu(II), DNA damage was not observed. Free nitrotyrosine induced slight DNA damage. NitroY-peptides damaged DNA more efficiently than free nitrotyrosine. The peptide containing three nitrotyrosine

Discussion

The present study has demonstrated that nitrotyrosine and nitroY-peptides of histone have an ability to cause Cu(II)-mediated DNA damage via the activation with P450 reductase. Inhibitory effects of catalase and bathocuproine suggested that H₂O₂ and Cu(I) were required for DNA damage. A possible mechanism of oxidative DNA damage induced by enzymatically activated nitrotyrosine can be speculated as accounting for most of the observations and references as follows. P450 reductase converts

Acknowledgements

This work was supported by a Grant-in-Aid from the Ministry of Education, Science, Sports and Culture of Japan.

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We also detected a decrease in 3-nitrotyrosine linked to the mature microbiome composition, which approximated levels detected in term infants (Figure 4E). This metabolite is an established marker of cell damage, inflammation, and nitric oxide production, and it is elevated in a large number of pathological inflammatory diseases (Murata and Kawanishi, 2004), including prematurity-related pathologies such as pulmonary dysplasia (Banks et al., 1998; Sheffield et al., 2006) and NEC (Egan et al., 2016), further supporting the benefits of microbiome maturation in the intervention group. L-cysteine, an important substrate for bifidobacteria (which are auxotroph for it; Ferrario et al., 2015), was reduced in the mature microbiomes (Figure 4E), which may reflect L-cysteine consumption by microbial communities with a greater Bifidobacterium abundance.
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The glomerular mesangial when activated imposes not only the thickening glomerular lining but also plays a crucial role in chronic renal inflammation [44]. NADPH oxidase-mediated 3-nitrotyrosine has also been studied in several inflammatory diseases [16,41,45,46]. To study the role of NOX2 mediated renal inflammation in this study, qRTPCR, and immunoreactivity of TLR4 and inflammatory cytokines were carried out.
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^☆: Abbreviations: nitroY-peptide, nitrotyrosine-containing peptide; RNS, reactive nitrogen species; 8-oxodG, 8-oxo-7,8-dihydro-2^′-deoxyguanosine (and also known as 8-hydroxy-2^′-deoxyguanosine); DTPA, diethylenetriamine-N,N,N^′,N″,N″-pentaacetic acid; HPLC-ECD, high performance liquid chromatography coupled with an electrochemical detector; NADPH, β-nicotinamide adenine dinucleotide phosphate (reduced form); P450 reductase, NADPH-cytochrome P450 reductase; SOD, superoxide dismutase.

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Oxidative DNA damage induced by nitrotyrosine, a biomarker of inflammation☆

Abstract

Section snippets

Materials and methods

Damage to 32P-labeled DNA fragments by nitrotyrosine in the presence of P450 reductase, NADPH, and Cu(II)

Discussion

Acknowledgements

Toxicol. Lett.

Arch. Biochem. Biophys.

Free Radic. Biol. Med.

Free Radic. Biol. Med.

Biochem. Biophys. Res. Commun.

Methods Enzymol.

Arch. Biochem. Biophys.

Methods Enzymol.

J. Biol. Chem.

J. Biol. Chem.

Methods Enzymol.

J. Biol. Chem.

Arch. Biochem. Biophys.

Free Radic. Biol. Med.

FEBS Lett.

Biochem. Biophys. Res. Commun.

Damage to ³²P-labeled DNA fragments by nitrotyrosine in the presence of P450 reductase, NADPH, and Cu(II)