Diurnal and day-to-day variation of urinary oxidative stress marker 8-hydroxy-2'-deoxyguanosine.

The urinary 8-hydroxy-2'-deoxyguanosine levels have been widely used as a biomarker of oxidative stress. The purpose of this study is to investigate the diurnal and day-to-day variations of urinary 8-hydroxy-2'-deoxyguanosine levels. For the diurnal variation, the urine samples were collected at the time of awakening and every 2 h, from 10:00 to 22:00, from 6 healthy participants. For the day-to-day variation, the urine samples were collected at the time of awakening for 35 consecutive days, from 27 healthy participants. As a result, no differences were observed in the diurnal urinary 8-hydroxy-2'-deoxyguanosine levels, and each subject had a characteristic 8-hydroxy-2'-deoxyguanosine level. On the other hand, the daily 8-hydroxy-2'-deoxyguanosine values showed a certain range of variation reflecting lifestyle factors, such as stress status, exercise, sleep time, drinking and diet. In conclusion, urinary 8-hydroxy-2'-deoxyguanosine may be a useful biomarker to control and prevent oxidative stress-related diseases, if the certain range of day-to-day variations of urinary 8-hydroxy-2'-deoxyguanosine is known. Even with only one measurement per year, the baseline urinary 8-hydroxy-2'-deoxyguanosine level could be achieved in a few years by incorporating the 8-hydroxy-2'-deoxyguanosine measurement as part of an annual health check. As the number of subjects was limited, further studies are needed for practical applications.

Pyrimidine Ring-Opened Product from Oxidative DNA Damage of 5-Formyl-2'-deoxyuridine.

After thymidine (dT) was treated with a Fenton-type reagent and further incubated for a long period (6 days) under physiological conditions (37 °C, pH 7.4), a new product, named dT*, was detected by HPLC in addition to the free thymine base and the known oxidative dT damage, 5-formyl-2'-deoxyuridine (f5dU). dT* was found to be formed from f5dU. The structure of dT* was determined to be 3-amino-2-carbamoyl-2-propenal-N3-2'-deoxyriboside, a pyrimidine ring-opened product from f5dU, on the basis of 1H- and 13C NMR analyses and mass spectra. From the model compound 1-methyl-5-formyluracil, a similar ring-opened product was formed after the incubation. dT* was also detected in DNA treated with a Fenton-type reagent or γ-rays, followed by the prolonged incubation. dT* will be a new promising marker of oxidative DNA damage. The possible role of this product in oxy-radical-induced mutagenesis is discussed.

Perceived Stress, Depressive Symptoms, and Oxidative DNA Damage.

OBJECTIVE: Psychosocial stress may influence the risk of disease through its association with oxidative DNA damage. We examined whether perceived stress and depressive symptoms were associated with urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), with mutual interaction on 8-OHdG. METHODS: This cross-sectional study included 6517 individuals aged 45 to 74 years who participated, between 2010 and 2012, in a follow-up survey of an ongoing cohort study. Perceived stress during the past year was measured using a self-report questionnaire. Depressive symptoms were evaluated using the Zung Self-Rating Depression Scale. Urinary 8-OHdG concentrations were measured using a column switching high-pressure liquid chromatography system coupled to an electrochemical detector. RESULTS: Higher perceived stress was significantly associated with higher 8-OHdG (2.1% increase per one-category increase of stress; ptrend = .025), even after adjusting for sex, age, supplement use, psychosocial factors, psychotropic medication use, smoking, and body mass index. This association was modestly attenuated after further adjustment for physical activity, suggesting possible mediation or confounding by this factor. Depressive symptoms were not significantly associated with 8-OHdG. No significant interaction was detected between perceived stress and depressive symptoms on 8-OHdG. CONCLUSIONS: In a general Japanese population, we found a weak positive association between perceived stress and urinary excretion of 8-OHdG, whereas no association was observed between depressive symptoms and 8-OHdG. Further studies are needed to examine whether the association between perceived stress and 8-OHdG is modified by depressive symptoms.

Intensity-specific effect of physical activity on urinary levels of 8-hydroxydeoxyguanosine in middle-aged Japanese.

Physical activity (PA) is recommended to both promote and maintain health and prevent cancer by improving the body's DNA repair system, which is considered a mechanism of cancer prevention. However, associations between PA and urinary levels of 8-hydroxydeoxyguanosine (8-OH-dG), which reflects DNA damage, are unclear. This cross-sectional study included 2370 men and 4052 women aged 45-74 years enrolled between 2010 and 2012. Habitual PA was assessed by single-axis accelerometer and urinary 8-OH-dG levels by automated HPLC. Multiple linear regression analysis was used to examine the relationship between log-transformed urinary 8-OH-dG and total PA (TPA) and PA of moderate/vigorous intensity (MVPA; ≥3 metabolic equivalents), with adjustment for age, body mass index, energy intake, alcohol consumption, smoking status, daily coffee drinking, menopause status (in women), and TPA (for MVPA). On multivariate adjustment, urinary 8-OH-dG levels were inversely correlated with TPA (ß = -0.020, P < 0.01) in women, and this correlation was not changed by PA intensity. Conversely, urinary 8-OH-dG levels were inversely correlated with MVPA (ß = -0.022, P < 0.05) in men, although the correlation with TPA was non-significant. This inverse correlation was clearer in current smokers than in never or former smokers, although the interaction between smoking status and MVPA on urinary 8-OH-dG levels was non-significant. In conclusion, greater TPA in women and greater MVPA in men were correlated with reduction in urinary 8-OH-dG, suggesting sex-specific effects of MVPA and TPA on protection from oxidative DNA damage. Increasing PA may mediate reduction in oxidative stress.

Coffee intake is associated with lower levels of oxidative DNA damage and decreasing body iron storage in healthy women.

Habitual coffee drinking has been linked to a lower risk for some forms of cancer, but the mechanism remains elusive. Coffee may decrease oxidative DNA damage, an important pathway to carcinogenesis. We examined the association between coffee consumption and urinary 8-hydroxydeoxyguanosine (8-OHdG) concentrations, a biomarker of systemic oxidative DNA damage and repair, in 507 healthy subjects (298 men and 209 women aged 21-67 yr) while adjusting for age, sex, smoking status, body mass index, job type, and fasting blood glucose in multivariable regression models. The association with green tea consumption was also assessed. Urinary 8-OHdG concentrations tended to decrease with coffee consumption in women (trend P = 0.046), with women drinking 2-3 cups of coffee per day showing the lowest mean of urinary 8-OHdG concentrations. This association was largely attenuated after further adjustment for serum ferritin concentrations, a marker of body iron storage (trend P = 0.96). Green tea consumption was not associated with urinary 8-OHdG concentrations. Coffee drinking may be associated with decreased systemic oxidative DNA damage through decreasing body iron storage in women.

Somatic cell mutations caused by 365 nm LED-UVA due to DNA double-strand breaks through oxidative damage.

Evidence is accumulating indicating that UVA (320-400 nm ultraviolet light) plays an important role in photo-carcinogenesis. UVA is thought to produce reactive oxygen species in irradiated cells through photo-activation of inherent photosensitizers, and was recently reported to cause DNA double-strand breaks (DSBs) in exposed cells. We have investigated the involvement of UVA in mutations and DNA damage in somatic cells using Drosophila melanogaster larvae. Using the Okazaki Large Spectrograph, we previously observed that longer wavelength UVA (>330 nm) was more mutagenic in post-replication repair-deficient D. melanogaster (mei-41) than in the nucleotide excision repair-deficient strain (mei-9). LED-light has recently been developed as a high-dose-rate UVA source. LED-UVA light (365 nm) was also more mutagenic in mei-41 than in mei-9. The mei-41 gene was shown to be an orthologue of the human ATR gene, which is involved in the repair of DSBs through phosphorylation of histone H2AX. In order to estimate the extent to which oxidative damage contributes to mutation, we established a new D. melanogaster strain (urate-null mutant) that is sensitive to oxidative damage and has a marker to detect somatic cell mutations. When somatic cell mutations were examined using this strain, LED-UVA was mutagenic in the urate-null strain at doses that were non-mutagenic in the urate-positive strain. In an effort to investigate the generation of DSBs, we examined the presence of phosphorylated histone H2AvD (H2AX D. melanogaster homologue). At high doses of LED-UVA (>800 kJ m(-2)), levels of phosphorylated H2AvD (γ-H2AvD) increased significantly in the urate-null strain. Moreover, the level of γ-H2AvD increased in the excision repair-deficient strain but not in the ATR-deficient strain following UVA-irradiation. These results supported the notion that the generation of γ-H2AvD was mediated by the function of the mei-41 gene. It was reported that ATR functions on DSB repair in D. melanogaster. Taken together, we propose a possible pathway for UVA-induced mutation, whereby DNA double-strand breaks resulting from oxidative stress might be responsible for UVA-induced mutation in somatic cells of D. melanogaster larvae.

Identification of octenal-related dA and dC adducts formed by reactions with a hemin-ω-6-fat peroxidation model system.

Deoxynucleosides were reacted in a lipid peroxidation model system, emulsified hemin-ethyl linoleate, and the adducts thus produced were analyzed by HPLC. Substantial amounts of stable adducts were detected in the dA- and dC-reaction mixtures. The structures of the major dA and dC adducts, other than the known 4-oxo-2-nonenal adducts, were determined to be etheno-type adducts, with a C6 side chain bearing an α-hydroxyl-group. These results suggested that the substance involved in adduct formation is 2,3-epoxyoctanal. This compound showed mutagenicity in Salmonella strains TA 100 and TA 104 without the S-9 mix. In addition, based on the structure of a minor dC adduct, another possibly involved mutagen, 4-oxo-2-octenal, was proposed. These mutagens may be formed during storage and cooking of food, or during digestion, and may be involved in human cancers.

Distributions and ranges of values of blood and urinary biomarker of inflammation and oxidative stress in the workers engaged in office machine manufactures: evaluation of reference values.

BACKGROUND: Interleukins, interferons and oxidative DNA products are important biomarkers assessing the inflammations and tissue damages caused by toxic materials in the body. We tried to evaluate distributions, reference values and age related changes of blood levels of inflammatory cytokines, C-reactive protein (CRP), IgE and urine levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) among workers in a cohort study evaluating the health influences of toner particles. METHODS: A total of 1366 male workers under age 50 years (age 19-49 years; 718 exposed and 648 not exposed to toner particles) in a cross sectional study of 1614 (categorized as 809 exposed and 805 not exposed, age 19-59 years) workers in a photocopier company has been followed prospectively as the cohort. Blood levels of interleukin (IL)-4, IL-6, IL-8, interferon-γ (IFN-γ), CRP, IgE and urine 8-OHdG were measured annually for 5 years. RESULTS: Reference values of the biomarkers are; CRP: 0.01-0.63×10(-2) g/L, IgE: 6-1480 IU/mL, IL-4: 2.6-76.1 pg/mL, IL-6: 0.4-4.9 pg/mL and 8-OHdG: 1.5-8.2 ng/mgCr. We could not evaluate reference values for IL-8 and IFN- γ because most of the values were below the sensitivity limits (2.0 pg/mL and 0.1 IU/mL, respectively). There were no differences of the biomarker levels between the toner exposed and the control workers. We observed a statistically significant age related decrease of serum IL-4 levels. CONCLUSIONS: This is the first report assessing the distributions and reference values of inflammatory biomarker levels in a large scaled cohort. We observed age related changes of some of the biomarkers. We could not detect any differences of the studied biomarker values between the toner exposed and the control workers.

Effects of Fe3O4 Magnetic Nanoparticles on A549 Cells.

Fe3O4 magnetic nanoparticles (MgNPs-Fe3O4) are widely used in medical applications, including magnetic resonance imaging, drug delivery, and in hyperthermia. However, the same properties that aid their utility in the clinic may potentially induce toxicity. Therefore, the purpose of this study was to investigate the cytotoxicity and genotoxicity of MgNPs-Fe3O4 in A549 human lung epithelial cells. MgNPs-Fe3O4 caused cell membrane damage, as assessed by the release of lactate dehydrogenase (LDH), only at a high concentration (100 µg/mL); a lower concentration (10 µg/mL) increased the production of reactive oxygen species, increased oxidative damage to DNA, and decreased the level of reduced glutathione. MgNPs-Fe3O4 caused a dose-dependent increase in the CD44+ fraction of A549 cells. MgNPs-Fe3O4 induced the expression of heme oxygenase-1 at a concentration of 1 µg/mL, and in a dose-dependent manner. Despite these effects, MgNPs-Fe3O4 had minimal effect on cell viability and elicited only a small increase in the number of cells undergoing apoptosis. Together, these data suggest that MgNPs-Fe3O4 exert little or no cytotoxicity until a high exposure level (100 µg/mL) is reached. This dissociation between elevated indices of cell damage and a small effect on cell viability warrants further study.

8-hydroxyguanine in urine and serum as an oxidative stress marker: effects of diabetes and aging.

8-Hydroxydeoxyguanosine (8-OH-dG) is the most extensively analyzed oxidative stress marker. Recently, 8-hydroxyguanine (free base: 8-OH-Gua) has been recognized as an oxidative stress marker. To verify the usefulness of 8-OH-Gua, the 8-OH-dG and 8-OH-Gua levels in the urine and the 8-OH-Gua levels in the serum of type 2 diabetic model animals, db/db mice, were measured as oxidative stress markers by a column switching HPLC-system coupled to an electrochemical detector. The urinary 8-OH-Gua and 8-OH-dG levels in db/db mice (7-26 weeks old) were significantly higher than those in control (db/m+) mice. The 8-OH-Gua levels in the serum of the db/db mice were also about 2-fold higher than those in the control mice at 26 weeks of age. In addition, the urinary levels of 8-OH-dG and 8-OH-Gua increased with age (9-26 weeks). A significant positive correlation was obtained between the 8-OH-dG and 8-OH-Gua levels in urine. Although no difference was observed in the 8-OH-dG levels in the liver and kidney DNA between the diabetic and control mice, these results suggested that urinary 8-OH-dG and free base 8-OH-Gua in urine or serum may be good biomarkers of oxidative stress.

Formation of the mutagenic DNA lesion 1,N2-ethenoguanine induced by heated cooking oil and identification of causative agents.

BACKGROUND: The DNA-damaging compounds in heated cooking oil were identified as guanosine adducts. Heated vegetable oil was subjected to deep-frying conditions at 170 °C for 45 min, reacted with isopropylidene guanosine (ipG) at pH 7.4, and the resulting compounds were separated by high-performance liquid chromatography (HPLC). RESULTS: Two adducts, 8-hydroxy-ipG and 1,N2-etheno-ipG, were identified in the reaction mixture. One of the major components in heated cooking oil, 2,4-heptadienal (HDE), efficiently produced etheno-ipG from ipG in the presence of tBuOOH. An oxidized HDE solution was fractionated using HPLC to identify causative agents, and each fraction was tested for etheno-ipG formation. In addition to the known lipid peroxidation product, 4,5-epoxy-2-heptenal, two unknown polar components with potent etheno-ipG formation activity were discovered. Based on Mass and UV spectra, their structures were identified as 6-oxo- and 6-hydroxy-2,4-HDE. Similarly, 6-oxo- and 6-hydroxy-2,4- decadienal (DDE) were formed from 2,4-DDE. Significant amounts of 6-oxo- and 6-hydroxy-2,4-alkadienal were detected in the heated cooking oil. These compounds induced the formation of 1,N2-ethenoguanine in nucleosides and DNA, especially in the presence of tBuOOH. Moreover, the formation of 6-oxo- and 6-OH-HDE from 2,4-HDE was accelerated in the presence of hemin and tBuOOH. CONCLUSION: The results suggest that these compounds are not only generated during the oil heating process but also produced from 2,4-alkadienal through digestion under normal physiological conditions, especially after ingesting heme- and alkyl-OOH-containing diets. Moreover, these compounds can be formed within cells under oxidative stress, potentially linking them to gastrointestinal carcinogenesis.

The Association between Oxidative Balance Score and Urinary Levels of 8-Hydroxydeoxyguanosine among Japanese Adults.

The oxidative balance score (OBS), wherein higher OBSs indicate lower oxidative stress, was designed to assess a composite measure of multiple pro-oxidant and antioxidant effects on an individual's oxidative stress status. This study aimed to evaluate whether OBSs were inversely associated with urinary levels of 8-hydroxydeoxyguanosine (8-OHdG)-an oxidative stress marker-among Japanese adults. This cross-sectional study was based on data obtained during 2010-2012. Overall, 7552 participants from the J-MICC Study Saga who answered a self-administered food frequency questionnaire were recruited for the final analysis. OBSs were calculated from 11 pro-oxidant and antioxidant lifestyle factors, including dietary intake, physical activity, and alcohol and smoking status. Urinary 8-OHdG values were corrected by creatinine level (ng/mg creatinine). Our findings revealed a higher total OBS was significantly associated with lower urinary 8-OHdG/creatinine levels after adjustment for covariates in men and women (p for trend < 0.01 in both sexes). We performed a multiple regression analysis of the association between OBSs and urinary 8-OHdG/creatinine levels stratified by age, body mass index (BMI), and menopausal status and found consistent negative associations in most groups for both sexes. No significant differences in the 60-64 age group for women (standardized ß = -0.09, p = 0.13) or BMI < 18.5 kg/m2 for men (standardized ß = -0.18, p = 0.17) were observed. A higher OBS had a strong inverse association with urinary 8-OHdG/creatinine levels in men and women among Japanese adults. The OBS might be a useful tool for evaluating the roles of oxidative stress-related lifestyle factors, including diet.

Inhibition by wheat sprout (Triticum aestivum) juice of bisphenol A-induced oxidative stress in young women.

For health of future generation, fertile young women should be monitored for exposure of endocrine disrupting chemicals (EDCs). Among EDCs, bisphenol A (BPA) is suggested to induce reactive oxygen species (ROS) which play an important role in pathologies of female diseases such as endometriosis. On the other hand, previous studies suggested that sprouts of wheat (Triticum aestivum) have antimutagenicity and antioxidant activity. We performed the 2 weeks intervention of wheat sprout juice (100ml/day) to investigate its effects on BPA-exposure and -oxidative toxicity in young women (N=14, age=24.4±4.0). Geometrical mean of urinary BPA levels was 1.81 (GSTD, 4.34)µg/g creatinine. We observed that irregular meals significantly increased levels of urinary BPA approximate 3 times (p=0.03). In addition, we found BPA-induced oxidative stress is correlated with levels of 8-hydroxydeoxyguanosine (8-OHdG) or malondialdehyde (MDA) levels (p=0.18 or 0.03, respectively). We also observed a continuous reduction of urinary BPA during the wheat sprout intervention (p=0.02). In summary, our data suggested potential detoxification of wheat sprouts on BPA-toxicity via antioxidative and interference of absorption, distribution, metabolism and excretion (ADME)-mediated mechanisms in young women.

New Plausible Mechanism for Gastric and Colorectal Carcinogenesis: Free Radical-Mediated Acetaldehyde Generation in a Heme/Myoglobin-Linoleate-Ethanol Mixture.

Epidemiological studies have revealed that alcohol, red meat, and cooking oil (or linoleate) are risk factors for both gastric and colon cancers. A survey of the mutation spectra of the p53 tumor suppressor gene in these cancers suggested that the types of mutations and the hot spots are similar to those induced by acetaldehyde (AcAld) in an in vitro p53 mutation analysis system. Accordingly, various combinations of possible factors, components, or model compounds were reacted in an emulsion and tested for the generation of AcAld. Efficient AcAld formation was only observed with combinations of three factors, red meat homogenate (or heme/myoglobin), methyl linoleate, and ethanol, but not by any combination of the two. The generated AcAld levels (ca. 500 µM) far exceeded the minimum mutagenic concentration (40-100 µM) obtained using concentrations of meat homogenate (or heme/Mb), linoleate, and ethanol comparable to those in the stomach after an ordinary meal. A mutagenic level of AcAld (75 µM) was also generated with a physiological concentration of ethanol, heme, and linoleate in the colon. As a mechanism, linoleate hydroperoxide formation and its decomposition in the presence of myoglobin (or heme) to generate the OH radical seem to be involved in the ethanol-to-AcAld conversion.

Free radical-mediated acetaldehyde formation by model reactions of dietary components: effects of meat, wine, cooking oil and coffee.

BACKGROUND: Alcohol consumption and the ingestion of red meat and oxidized cooking oil are risk factors of gastric and colorectal cancers. We reported that acetaldehyde (AcAld) is generated from Heme/Mb/Meat-Linoleate-EtOH model reaction mixtures, and thus could be a new plausible mechanism for the carcinogenesis (Kasai and Kawai, ACS Omega, 2021). RESULTS: In this study, we investigated the effects of wine and coffee, in addition to meat components, on this reaction. Depending on the conditions, such as pH, reaction time and choice of free hemin, myoglobin (Mb), as well as meat extracts (raw meat, baked meat, salami), wine and coffee enhanced AcAld formation. Polyphenols in red wine and coffee may stimulate AcAld formation by acting as pro-oxidants in the presence of Heme/Mb/Meat. In a model reaction of Mb + EtOH + H2O2, we observed time-dependent AcAld formation. In support of these in vitro data, after the consumption of a red meat-rich diet with red wine, the fecal AcAld level significantly increased as compared to the levels associated with a diet of fish + wine, or red meat without alcohol. CONCLUSIONS: These results suggested that AcAld generation from dietary components may be an important mechanism of gastrointestinal tract carcinogenesis.

Involvement of specialized DNA polymerases in mutagenesis by 8-hydroxy-dGTP in human cells.

The mutagenicity of an oxidized form of dGTP, 8-hydroxy-2'-deoxyguanosine 5'-triphosphate (8-OH-dGTP), was examined using human 293T cells. Shuttle plasmid DNA containing the supF gene was first transfected into the cells, and then 8-OH-dGTP was introduced by means of osmotic pressure. The DNAs replicated in the cells were recovered and then transfected into Escherichia coli. 8-OH-dGTP induced A:T-->C:G substitution mutations in the cells. The knock-downs of DNA polymerases eta and zeta, and REV1 by siRNAs reduced the A:T-->C:G substitution mutations, suggesting that these DNA polymerases are involved in the misincorporation of 8-OH-dGTP opposite A in human cells. In contrast, the knock-down of DNA polymerase iota did not affect the 8-OH-dGTP-induced mutations. The decrease in the induced mutation frequency was more evident by double knock-downs of DNA pols eta plus zeta and REV1 plus DNA pol zeta (but not by that of DNA pol eta plus REV1), suggesting that REV1-DNA pol eta and DNA pol zeta work in different steps. These results indicate that specialized DNA polymerases are involved in the mutagenesis induced by the oxidized dGTP.

Body iron store as a predictor of oxidative DNA damage in healthy men and women.

While iron plays an important role in many cellular functions, excess iron storage induces DNA damage by generating hydroxyl radicals and thus promotes carcinogenesis. However, it remains unclear whether body iron levels that are commonly observed in a general population are related to oxidative DNA damage. We examined the association between serum ferritin concentrations and levels of urinary 8-hydroxydeoxyguanosine (8-OHdG), a biomarker of systemic oxidative DNA damage and repair, in 528 Japanese men and women aged 21-67 years. Men had much higher ferritin levels than in women, and the levels were significantly greater in women aged 50 years or older than in women aged less than 50 years. Urinary 8-OHdG concentrations were significantly and positively associated with serum ferritin levels in all the subgroups. The Spearman rank correlation coefficients were 0.47, 0.76, and 0.73 for men overall, women aged less than 50 years, and women aged 50 years or older, respectively. These associations were materially unchanged after adjustment for potential confounding variables. In men, a more pronounced association was observed in nonsmokers than in smokers. Our results suggest body iron storage is a strong determinant of levels of systemic oxidative DNA damage in a healthy population.

Detection of lipid peroxidation-induced DNA adducts caused by 4-oxo-2(E)-nonenal and 4-oxo-2(E)-hexenal in human autopsy tissues.

DNA adducts are produced both exogenously and endogenously via exposure to various DNA-damaging agents. Two lipid peroxidation (LPO) products, 4-oxo-2(E)-nonenal (4-ONE) and 4-oxo-2(E)-hexenal (4-OHE), induce substituted etheno-DNA adducts in cells and chemically treated animals, but the adduct levels in humans have never been reported. It is important to investigate the occurrence of 4-ONE- and 4-OHE-derived DNA adducts in humans to further understand their potential impact on human health. In this study, we conducted DNA adductome analysis of several human specimens of pulmonary DNA as well as various LPO-induced DNA adducts in 68 human autopsy tissues, including colon, heart, kidney, liver, lung, pancreas, small intestine, and spleen, by liquid chromatography tandem mass spectrometry. In the adductome analysis, DNA adducts derived from 4-ONE and 4-OHE, namely, heptanone-etheno-2'-deoxycytidine (HεdC), heptanone-etheno-2'-deoxyadenosine (HεdA), and butanone-etheno-2'-deoxycytidine (BεdC), were identified as major adducts in one human pulmonary DNA. Quantitative analysis revealed 4-ONE-derived HεdC, HεdA, and heptanone-etheno-2'-deoxyguanosine (HεdG) to be ubiquitous in various human tissues at median values of 10, 15, and 8.6 adducts per 10(8) bases, respectively. More importantly, an extremely high level (more than 100 per 10(8) bases) of these DNA adducts was observed in several cases. The level of 4-OHE-derived BεdC was highly correlated with that of HεdC (R(2) = 0.94), although BεdC was present at about a 7-fold lower concentration than HεdC. These results suggest that 4-ONE- and 4-OHE-derived DNA adducts are likely to be significant DNA adducts in human tissues, with potential for deleterious effects on human health.

DNA modifications by the omega-3 lipid peroxidation-derived mutagen 4-oxo-2-hexenal in vitro and their analysis in mouse and human DNA.

4-Oxo-2-hexenal (4-OHE), which forms a 2'-deoxyguanosine (dG) adduct in a model lipid peroxidation system, is mutagenic in the Ames test. It is generated by the oxidation of omega-3 fatty acids and is commonly found in dietary fats, such as fish oil, perilla oil, rapeseed oil, and soybean oil. 4-OHE also forms adducts with 2'-deoxyadenosine (dA), 2'-deoxycytidine (dC), and 5-methyl-2'-deoxycytidine (5-Me-dC) in DNA. In this study, we characterized the structures of these adducts in detail. We measured the amounts of 4-OHE-DNA adducts in mouse organs by LC/MS/MS, after 4-OHE was orally administered to mice. The 4-OHE-dA, 4-OHE-dC, 4-OHE-dG, and 4-OHE-5-Me-dC adducts were detected in stomach and intestinal DNA in the range of 0.25-43.71/10(8) bases. After the 4-OHE administration, the amounts of these DNA adducts decreased gradually over 7 days. We also detected 4-OHE-dC in human lung DNA, in the range of 2.6-5.9/10(9) bases. No difference in the 4-OHE adduct levels was detected between smokers and nonsmokers. Our results suggest that 4-OHE-DNA adducts are formed by endogenous as well as environmental lipid peroxides.

DNA methylation by dimethyl sulfoxide and methionine sulfoxide triggered by hydroxyl radical and implications for epigenetic modifications.

In this Letter, we demonstrate the formation of m(5)dC from dC or in DNA by dimethylsulfoxide (DMSO) and methionine sulfoxide (MetO), under physiological conditions in the presence of the Fenton reagent in vitro. DMSO reportedly affects the cellular epigenetic profile, and enhances the metastatic potential of cultured epithelial cells. The methionine sulfoxide reductase (Msr) gene was suggested to be a metastatis suppressor gene, and the accumulation of MetO in proteins may induce metastatic cancer. Our findings are compatible with these biological data and support the hypothesis that chemical cytosine methylation via methyl radicals is one of the mechanisms of DNA hypermethylation during carcinogenesis. In addition to m(5)dC, the formation of 8-methyldeoxyguanosine (m(8)dG) was also detected in DNA under the same reaction conditions. The m(8)dG level in human DNA may be a useful indicator of DNA methylation by radical mechanisms.