8-hydroxyguanine and 8-hydroxyguanosine

Low birthweight (LBW) has been associated with an increased risk of development of type 2 diabetes in adult life. Both type 1 and type 2 diabetes mellitus are characterized by increased oxidative stress. The purpose of this study was to investigate whether young healthy adults born with LBW showed differences in oxidative stress under normal conditions and during the added challenge of a physiological Intralipid infusion.. Urinary excretion of DNA markers of oxidative stress were analyzed by LC-MS/MS in 19 men (aged 19 years) with LBW and in 19 age matched, normal birthweight (NBW) controls pre- and post a 3-fold increase of plasma free fatty acids.. Mean excretion rates of 8-oxo-guanine (8oxoGua), 8-oxo-guanosine (8oxoGuo), 8-oxo-2'deoxyguanosine (8oxodG), and 1,N6-ethenodeoxyadenosine (epsilon dA) did not statistically differ between subjects with LBW and NBW (66.9 versus 73.9 nmol/15 h, 17.8 versus 18.5 nmol/15 h, 11.9 versus 14.4 nmol/15 h and 44.0 versus 43.2 pmol/15 h, respectively). Furthermore, Intralipid infusion did not affect excretion of DNA adducts in LBW or NBW subjects. Statistically significant correlations were found between body mass index and urinary excretion of 8oxoGua (r = 0.64, p = 0.003) and 8oxoGuo (r = 0.64, p = 0.003) in the LBW group only.. These findings suggest that oxidative stress may be a consequence of diabetes and is not, or at least only partly, involved in the early pathogenesis of type 2 diabetes.

Topics: Adult; Biomarkers; Birth Weight; Body Mass Index; DNA; Fatty Acids, Nonesterified; Guanine; Guanosine; Humans; Infant, Low Birth Weight; Infant, Newborn; Male; Oxidative Stress

One study found higher leukocytes 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in patients with spontaneous intracerebral hemorrhage (ICH) than in healthy subjects due to the oxidation of guanosine from deoxyribonucleic acid (DNA). The objective of this study was to determine whether there is an association between oxidative damage of serum DNA and ribonucleic acid (RNA) and mortality in patients with ICH.. In this observational and prospective study, patients with severe supratentorial ICH (defined as Glasgow Coma Scale < 9) were included from six Intensive Care Units of Spanish hospitals. At the time of severe ICH diagnosis, concentrations in serum of malondialdehyde (as lipid peroxidation biomarker) and of the three oxidized guanine species (OGS) (8-hydroxyguanosine from RNA, 8-hydroxyguanine from DNA or RNA, and 8-OHdG from DNA) were determined. Thirty-day mortality was considered the end-point study.. Serum levels of OGS (p < 0.001) and malondialdehyde (p = 0.002) were higher in non-surviving (n = 46) than in surviving patients (n = 54). There was an association of serum OGS levels with serum malondialdehyde levels (rho = 0.36; p = 0.001) and 30-day mortality (OR = 1.568; 95% CI 1.183-2.078; p = 0.002).. The novel and most important finding of our study was that serum OGS levels in ICH patients are associated with mortality.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aged; Cerebral Hemorrhage; DNA; DNA Damage; Female; Glasgow Coma Scale; Guanine; Guanosine; Humans; Male; Middle Aged; Mortality; Oxidative Stress; Prognosis; Prospective Studies; RNA

The hyperoxidative state in traumatic brain injury (TBI) could produce oxidative damage on the ribonucleic acid (RNA) and deoxyribonucleic acid (DNA). Oxidative damage to nucleic acids in TBI patients has been studied, and higher concentrations of 8-OHdG were found in postmortem brain samples of subjects who died following TBI than in subjects who died from sudden cardiac death. Thus, the objective of this study was to determine whether there is an association between serum DNA and RNA oxidative damage and mortality in TBI patients.. We included patients with severe isolated TBI defined as a lower score than 9 points in the Glasgow Coma Scale (GCS) and lower than 9 points in non-cranial aspects in the Injury Severity Score. We determined serum concentrations of the three oxidized guanine species (OGS) (8-OHdG from DNA, 8-hydroxyguanosine from RNA, and 8-hydroxyguanine from DNA or RNA) and malondialdehyde (to estimate lipid peroxidation) on the day of TBI. Mortality at 30 days was the end-point study.. We found higher serum concentrations of OGS (p < 0.001) and malondialdehyde (p < 0.001) in non-surviving (n = 34) than in surviving patients (n = 90), an association between serum OGS levels and 30-day mortality after control for CGS, age, and computed tomography findings (OR = 1.397; 95% CI = 1.137-1.716; p = 0.001), and a positive correlation between serum levels of OGS and malondialdehyde (rho = 0.24; p = 0.01).. To our knowledge, our study is the largest series reporting data on DNA oxidative damage in TBI patients and is the first reporting DNA and RNA oxidative damage in TBI patients associating lipid peroxidation and mortality.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Aged; Brain Injuries, Traumatic; DNA Damage; Female; Glasgow Coma Scale; Guanine; Guanosine; Humans; Injury Severity Score; Lipid Peroxidation; Male; Malondialdehyde; Middle Aged; Mortality; Odds Ratio; Oxidative Stress; RNA

A reliable and fast liquid chromatography-tandem mass spectrometry method has been developed for the simultaneous determination of three oxidized nucleic acid damage products in urine, 8-oxoguanine (8-oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and 8-oxo-7,8-dihydroguanosine (8-oxoGuo). We applied this method to assess the effect of various urine workup procedures on the urinary concentrations of the oxidized nucleic acid products. Our results showed that frozen urine samples must be warmed (i.e., to 37 °C) to re-dissolve any precipitates prior to analysis. We showed that common workup procedures, such as thawing at room temperature or dilution with deionized water, are not capable of releasing fully the oxidized nucleic acid products from the precipitates, and result in significant underestimation (up to ~ 100% for 8-oxoGua, ~ 86% for both 8-oxodGuo and 8-oxoGuo). With this method, we further assessed and compared the ability of the three oxidized nucleic acid products, as well as malondialdehyde (MDA, a product of lipid peroxidation), to biomonitor oxidative stress in vivo. We measured a total of 315 urine samples from subjects with burdens of oxidative stress from low to high, including healthy subjects, patients with chronic obstructive pulmonary disease (COPD), and patients on mechanical ventilation (MV). The results showed that both the MV and COPD patients had significantly higher urinary levels of 8-oxoGua, 8-oxodGuo, and 8-oxoGuo (P < 0.001), but lower MDA levels, compared to healthy controls. Receiver operating characteristic curve analysis revealed that urinary 8-oxoGuo is the most sensitive biomarker for oxidative stress with area under the curve (AUC) of 0.91, followed by 8-oxodGuo (AUC: 0.80) and 8-oxoGua (AUC: 0.76). Interestingly, MDA with AUC of 0.34 failed to discriminate the patients from healthy controls. Emerging evidence suggests a potential clinical utility for the measurement of urinary 8-oxoGuo, and to a lesser extent 8-oxodGuo, which is strongly supported by our findings.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aged; Biomarkers; Chromatography, Liquid; Deoxyguanosine; Female; Guanine; Guanosine; Humans; Male; Metabolome; Metabolomics; Middle Aged; Oxidative Stress; Pulmonary Disease, Chronic Obstructive; Reactive Oxygen Species; ROC Curve; Tandem Mass Spectrometry; Temperature

3-hydroxy-3-methylglutaric aciduria (HMGA) is an inherited disorder of the leucine catabolic pathway in which occurs a deficiency of the 3-hydroxy-3-methylglutaryl-CoA lyase enzyme. Therefore, the organic acids 3-hydroxy-3-methylglutaric (HMG) and 3-methylglutaric (MGA), mainly, accumulate in tissues of affected patients. Lately, much attention has been focused on free radicals as mediators of tissue damage in human diseases, causing lipid peroxidation, protein oxidation and DNA damage. The treatment of this disease is based in a restricted protein ingest and supplementation with l-carnitine (LC), an antioxidant and detoxifying agent. In the present work, we investigated the in vitro oxidative damage to DNA induced by the accumulation of organic acids and oxidative stress parameters in vivo of patients with 3-HMG, as well as the effect of the recommended therapy. The in vitro DNA damage was analyzed by the alkaline comet assay in leukocytes incubated with HMG and MGA (1 mM, 2.5 mM and 5 mM) and co-incubated with LC (90 μM and 150 μM). The in vivo urinary 15-F2t-isoprostane levels and urinary oxidized guanine species were measured by ELISA kits in patient's urine before and after the treatment with LC. HMG and MGA induced a DNA damage index (DI) significantly higher than that of the control group. The DI was significantly reduced in the presence of LC. It was also verified a significant increase of oxidized guanine species and urinary isoprostane levels, biomarker of oxidative DNA damage and lipid peroxidation respectively, in patients before treatment. After the treatment and supplementation with LC, patients presented significantly lower levels of those biomarkers. Analyzing the data together, we can conclude that HMGA patients present oxidative lipid and DNA damage, which is induced by HMG and MGA, and the antioxidant therapy with LC can prevent that kind of injuries.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acetyl-CoA C-Acetyltransferase; Adolescent; Amino Acid Metabolism, Inborn Errors; Carnitine; Child; Child, Preschool; Dinoprost; DNA Damage; Guanine; Guanosine; Humans; Infant; Lipid Peroxidation; Meglutol

This study aimed to identify sensitive and not-invasive biomarkers of early genotoxic/oxidative effect for exposure to styrene in the fibreglass reinforced plastic manufacture. We studied 11 workers of a plastic manufacture using open molding process (A), 16 workers of a manufacture using closed process (B) and 12 controls. We evaluated geno/cytotoxic effects on buccal cells by Buccal Micronucleus Cytome (BMCyt) assay and genotoxic/oxidative effects on lymphocytes by Fpg-comet test. On A workers we also evaluated urinary 8oxoGua, 8oxodGuo and 8oxoGuo to investigate oxidative stress. Personal inhalation exposure to styrene was monitored by passive air sampling and GC/MS. Biological monitoring included urinary metabolites mandelic acid (MA) and phenylglyoxylic acid (PGA). The findings show higher styrene exposure, urinary MA + PGA levels and micronucleus frequency in manufacture A. Higher buccal karyolytic cell frequency vs controls were found in both exposed populations. We found in exposed workers, no induction of direct DNA damage but oxidative DNA damage. Fpg-comet assay and urinary oxidized guanine seem to be sensitive biomarkers of oxidative stress and BMCyt assay a good-not invasive biomarker of cyto-genotoxicity at target organ. The study, although limited by the small number of studied subjects, shows the usefulness of used biomarkers in risk assessment of styrene-exposed workers.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Air Pollutants, Occupational; Case-Control Studies; Comet Assay; Deoxyguanosine; DNA Damage; Environmental Biomarkers; Environmental Monitoring; Female; Glass; Guanine; Guanosine; Humans; Inhalation Exposure; Lymphocytes; Male; Manufacturing Industry; Micronuclei, Chromosome-Defective; Micronucleus Tests; Middle Aged; Mouth Mucosa; Occupational Exposure; Occupational Health; Oxidative Stress; Pilot Projects; Reproducibility of Results; Risk Assessment; Styrene; Urinalysis

Chemotherapy is less often prescribed in older individuals due to concerns about post-treatment morbidity and quality of life. We evaluated the physical performance of breast cancer survivors treated with and without adjuvant chemotherapy.. We conducted a case-control study in 56 estrogen receptor positive breast cancer survivors (BCS) on adjuvant aromatase inhibitors 1-2years after definitive surgery. Cases had received adjuvant chemotherapy (n=27; age 70.5±3.6years) versus age-matched controls who had not (n=29; age 70.0±4.3years). Measures of grip strength, physical activity and performance, walking speed, fatigue, and self-reported physical function were collected. Biological correlates of inflammation, frailty and markers of DNA and RNA oxidation were compared.. Grip strength (controls: 21±7.4 vs.. 29.7±5.0kg, p=0.20), physical activity (5403±3204 vs. 6801±9320steps/day, p=0.45), physical performance (short physical performance battery score: 10.1±1.8 vs. 10.4±1.1, p=0.52) and long-distance walking speed (1.2±0.21 vs. 1.3±0.41m/s, p=0.17) were similar between the two groups. Self-reported physical function was marginally lower in cases than controls (controls: 72±24 vs.. 57±34AU, p=0.07). Fatigue disruptiveness was not different between groups (controls: 11.1±13.0 vs.. 15.7±16.2AU, p=0.24). Similarly, the inflammation, oxidation, and frailty markers did not present a significant difference between groups, except for vitamin D levels (p=0.04).. Older women who received chemotherapy reported having slightly lower physical function, but a similar physical performance compared to women who did not. These data suggest that older BCS treated with chemotherapy recover to an extent similar to survivors who only received hormonal therapy.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Activities of Daily Living; Aged; Aromatase Inhibitors; Biomarkers; Breast Neoplasms; Cancer Survivors; Case-Control Studies; Chemotherapy, Adjuvant; Cross-Sectional Studies; Deoxyguanosine; Exercise; Fatigue; Female; Fibrin Fibrinogen Degradation Products; Guanine; Guanosine; Hand Strength; Humans; Insulin-Like Growth Factor Binding Protein 3; Insulin-Like Growth Factor I; Interleukin-6; Oxidation-Reduction; Pyrimidines; Serum Albumin; Tandem Mass Spectrometry; Tumor Necrosis Factor-alpha; Vitamin D; Walking Speed

Evaluating the correlation between otoacoustic emission levels, styrene exposure, and oxidative stress biomarkers concentration in styrene-exposed subjects, to investigate the role of oxidative stress in outer hair cell damage.. Distortion product otoacoustic emissions were measured in the exposed workers and in a control group. Separation between the distortion and reflection otoacoustic components was performed by time-frequency-domain filtering. The urinary concentration of the DNA and RNA oxidation products, namely 8-oxo-7,8-dihydroguanine (oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (oxodGuo), and 8-oxo-7,8-dihydroguanosine (oxoGuo), were evaluated.. Nine subjects exposed to styrene in a fiberglass factory, eight control subjects. The two groups were statistically equivalent in mean age.. Statistically significant differences were found in the distortion component levels between the exposed and the control group. High levels of the oxidative damage biomarkers were found in the workers exposed to high levels of styrene. Significant negative correlation was found between the otoacoustic emission distortion component levels and the concentration of the oxoGuo biomarker.. Exposure-induced damage of the cochlear amplifier is shown in the mid-frequency range, confirming animal experiments, in which hair cells in the cochlear middle turn were damaged. Hearing damage is consistent with the outer hair cell apoptosis pathway associated with oxidative stress.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acoustics; Adult; Apoptosis; Biomarkers; Case-Control Studies; Deoxyguanosine; DNA Damage; Female; Guanine; Guanosine; Hair Cells, Auditory, Outer; Hearing Loss, Noise-Induced; Hearing Tests; Humans; Male; Middle Aged; Noise, Occupational; Occupational Diseases; Occupational Exposure; Occupational Health; Otoacoustic Emissions, Spontaneous; Oxidative Stress; Risk Factors; Styrene

The oxidized nucleoside 8-hydroxy-2'-deoxyguanosine has been widely studied as a marker of DNA oxidation; however, data on the occurrence of other metabolites in plasma that are related to DNA damage are scarce. We have applied an improved, sensitive, robust, and reliable method, involving solid phase extraction and ultrahigh-performance liquid chromatography (UHPLC)-tandem mass spectrometry (MS/MS), to the precise quantitation of seven metabolites in the plasma of 15 elite triathletes after a 2-week training program. All compounds were eluted in the first 1.6 min, with limits of detection and quantification ranging between 0.001 and 0.3 ng.mL(-1) and 0.009 and 0.6 ng.mL(-1), respectively. Four compounds were detected in plasma: guanosine-3'-5'-cyclic monophosphate, 8-hydroxyguanine, 8-hydroxy-2'-deoxyguanosine, and 8-nitroguanosine. After two weeks of training, 8-hydroxyguanine exhibited the highest increase (from 0.031 ± 0.008 nM to 0.036 ± 0.012 nM) (p < 0.05), which could be related to the enhanced activity of DNA-repairing enzymes that excise this oxidized base. Increased levels of guanosine-3'-5'-cyclic monophosphate and 8-hydroxy-2'-deoxyguanosine were also observed. In contrast, levels of 8-nitroguanosine (p < 0.05) were significantly reduced, which might be a protective measure as this compound strongly stimulates the generation of superoxide radicals, and its excess is related to pathologies such as microbial (viral) infections and other inflammatory and degenerative disorders. The results obtained indicate an induced adaptive response to the increased oxidative stress related to elite training, and point to the benefits associated with regular exercise.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Athletes; Cyclic GMP; Deoxyguanosine; DNA; DNA Fragmentation; Female; Guanine; Guanosine; Humans; Limit of Detection; Male; Nitro Compounds; Oxidation-Reduction; Oxidative Stress; Physical Conditioning, Human; Young Adult

The aim of this work was to evaluate the oxidative damage to nucleic acids in children (5-11 years) associated with exposure to environmental pollutants and tobacco smoke (ETS). For each subject, urinary sampling was done twice (evening and next morning) to measure by tandem LC-MS-MS such oxidated products of nucleic acids as 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanosine (8-oxoGuo), and 8-oxo-7,8-dihydroguanine (8-oxoGua). Methyl tert-butyl ether (U-MTBE), benzene (U-Benz), and its metabolites (t,t-muconic and S-phenylmercapturic acids, t,t-MA and S-PMA, respectively) were determined as biomarkers of exposure to air pollution, and cotinine as a biomarker of exposure to ETS. Biomarkers of exposure (S-PMA and U-MTBE) and of DNA oxidation (8-oxodGuo) were dependent on the urbanization and industrialization levels and increased in the evening sample as compared to next morning (p<0.05). In both evening and next morning samples, 8-oxodGuo and 8-oxoGuo correlated with each other (r=0.596 and r=0.537, respectively, p<0.01) and with biomarkers of benzene exposure, particularly S-PMA (r=0.59 and r=0.45 for 8-oxodGuo and r=0.411 and r=0.383 for 8-oxoGuo, p<0.01). No such correlations were observed for U-MTBE and cotinine. Multiple linear regression analyses showed that 8-oxodGuo was positively associated with S-PMA at both sampling times (β=0.18 and β=0.14 for evening and next morning sampling, respectively; p<0.02) and weakly with U-MTBE (β=0.07, p=0.020) only in the evening urines. These results suggest that the selected biomarkers of exposure to benzene, particularly S-PMA, are good tracers of exposure to complex mixtures of oxidative pollutants and that the associated oxidative damage to nucleic acids is detectable even at very low levels of exposure.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acetylcysteine; Air Pollutants; Benzene; Biomarkers; Child; Child, Preschool; Cotinine; Deoxyguanosine; DNA; DNA Damage; Environmental Monitoring; Female; Guanine; Guanosine; Humans; Male; Methyl Ethers; Oxidation-Reduction; Sicily

Subjects with chronic renal failure (CRF) exhibit oxidative genome damage, which may predispose to carcinogenesis, and Gum acacia (GumA) ameliorates this condition in humans and animals. We evaluated here renal DNA damage and urinary excretion of four nucleic acid oxidation adducts namely 8-oxoguanine (8-oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), 8-oxoguanosine (8-oxoGuo) and 8-hydroxy-2-deoxyguanisone (8-OHdg) in rats with adenine (ADE)-induced CRF with and without GumA treatment.. Twenty-four rats were divided into four equal groups and treated for 4 weeks. The first group was given normal food and water (control). The second group was given normal food and GumA (15% w/v) in drinking water. The third group was fed powder diet containing adenine (ADE) (0·75% w/w in feed). The fourth group was fed like in the third group, plus GumA in drinking water (15%, w/v).. ADE feeding induced CRF (as measured by several physiological, biochemical and histological indices) and also caused a significant genetic damage and significant decreases in urinary 8-oxo Gua and 8-oxoGuo, but not in the other nucleic acids. However, concomitant GumA treatment reduced the level of genetic damage in kidney cells as detected by Comet assay and significantly reversed the effect of adenine on urinary 8-oxoGuo.. Treatment with GumA is able to mitigate genetic damage in renal tissues of rats with ADE-induced CRF.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adenine; Animals; Comet Assay; Deoxyguanosine; DNA Damage; Guanine; Guanosine; Gum Arabic; Kidney Failure, Chronic; Kidney Function Tests; Male; Random Allocation; Rats, Wistar; Renal Agents

The aims of the present study were to establish the background levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanosine (8-oxoGuo), 8-oxo-7,8-dihydroguanine (8-oxoGua) among a group of healthy Italian children, and to evaluate the contribution of some potential interfering/confounding factors to the urinary (u) levels of these biomarkers of oxidised guanine derivatives. The levels of 8-oxodGuo, 8-oxoGuo, 8-oxoGua, and u-cotinine in urine samples from 159 healthy children (5-11years) recruited in a cross-sectional study were measured via liquid chromatography-tandem mass spectrometry. Data regarding the anthropometric and life-style characteristics of the participants were obtained from questionnaires. The 5th-95th percentiles of the levels of 8-oxodGuo, 8-oxoGuo, and 8-oxoGua for all children were 2.4-13.9, 3.8-19.9 and 5.4-79.5μg/L and 2.9-12.6, 4.8-15.2, and 5.1-93.4μg/g creatinine, respectively. Significant correlations were found between the level of 8-oxoGuo and that of 8-oxoGua and 8-oxodGuo but not between the level of 8-oxoGua and that of 8-oxodGuo in all children and in both the male and female subgroups. Multiple linear regression analyses revealed the independent effect of the investigated variables on 8-oxodGuo, 8-oxoGuo, and 8-oxoGua. u-Creatinine was the most significant predictor of the urinary excretion of both 8-oxoGuo and 8-oxodGuo, age displayed a significant positive independent effect on the level of 8-oxoGuo, whereas the weight status according to the BMI was negatively associated with the level of 8-oxodGuo. None of the chosen independent variables influenced the levels of 8-oxoGua.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Biomarkers; Body Mass Index; Child; Cotinine; Cross-Sectional Studies; Deoxyguanosine; Environmental Exposure; Female; Guanine; Guanosine; Humans; Life Style; Male; Oxidative Stress; Urban Population

This study was aimed at defining the reference ranges for biomarkers of oxidized guanine in (2'-deoxy)ribonucleotides and nucleic acids from a large Italian sample. We recruited 300 healthy subjects (150 males; mean age 44.1±13.6years; 26% smokers) without any known exposure to occupational oxidizing agents. They were asked to provide a spot urine sample, on which the following markers were determined by liquid chromatography-tandem mass spectrometry: 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanosine (8-oxoGuo), 8-oxo-7,8-dihydroguanine (8-oxoGua), and cotinine. The reference ranges, estimated as the 5th-95th percentiles of creatinine-normalized values (pmol/μmol(creat)) were 0.7-4.2, 0.9-4.7, and 5.6-120.7 for 8-oxodGuo, 8-oxoGuo, and 8-oxoGua, respectively. Oxidation biomarkers were correlated with one another (p<0.005) and with urinary creatinine (p<0.0001). Males excreted significantly higher concentrations of 8-oxoGua than females (p<0.0001). 8-OxoGua and 8-oxoGuo showed a positive association with age (p<0.001), also after stratification by gender. Multiple linear regression models including urinary creatinine concentration, age, and smoking habit as independent variables showed a significant effect of age, but not of smoking, on the levels of 8-oxoGuo in males (p<0.0001) and of both 8-oxoGuo and 8-oxoGua in females (p<0.0001). A preliminary assessment in a small group (n=25) of patients affected by advanced non-small-cell lung cancer and receiving platinum-based chemotherapy showed significantly higher values of both 8-oxoGuo and 8-oxodGuo (p<0.0001 for both) compared to the referent population.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Age Factors; Aged; Aged, 80 and over; Biomarkers; Carcinoma, Non-Small-Cell Lung; Chromatography, Liquid; Cotinine; Deoxyguanosine; Female; Guanine; Guanosine; Humans; Lung Neoplasms; Male; Middle Aged; Nucleic Acids; Oxidation-Reduction; Oxygen; Reference Values; Smoking; Tandem Mass Spectrometry; Young Adult

We developed a new method for the simultaneous quantitative determination of 8-oxo-7,8-hydro-2'-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanine (8-oxoGua), 8-oxo-7,8-dihydroguanosine (8-oxoGuo), and the corresponding non-oxidized forms, 2'-deoxyguanosine (dGuo), guanine (Gua) and guanosine (Guo), in human urine samples by liquid chromatography-tandem mass spectrometry. Differences in the ionization of analytes in different urine batches with variable matrix effects were effectively compensated for by internal standardization with stable isotope-labelled analytes. The method was sensitive enough to allow the determination of background levels of these biomarkers and was applied to characterize the inter- and intraindividual variability of biomarkers in the diurnal profile of concentrations in 24 healthy volunteers. When normalized for creatinine, none of the biomarkers was affected by sampling time, thus ruling out any circadian rhythm for nucleic acid oxidation in urine.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Biomarkers; Chromatography, Liquid; Circadian Rhythm; Creatinine; Deoxyguanosine; Guanine; Guanosine; Humans; Isotopes; Mass Spectrometry; Nucleic Acids; Oxygen; Reference Values; Reproducibility of Results

A sensitive and specific assay aimed at measuring the oxidized nucleic acids, 8-oxoguanine (8-oxoGua), fapy-guanine (Fapy-Gua), 8-oxoguanosine (8-oxoGuo), 8-oxo-2'-deoxyguanosine (8-oxodG) has been developed by coupling reversed phase liquid chromatography (HPLC) with electrospray tandem mass spectrometry detection (MS/MS) and isotope dilution. The HPLC-MS/MS approach with multiple reaction monitoring (MRM) allowed for the sensitive determination of 8-oxoGua, Fapy-Gua, 8-oxoGuo, and 8-oxodG in human urine samples. There is no sample preparation needed except for the addition of buffer and (13)C- and (15)N-labeled internal standards to the urine prior to sample injection into the HPLC-MS/MS system. This method was tested in urine samples from non-smokers, smokers, non-smokers with chronic kidney disease (CKD) and smokers with CKD, to assess the level of oxidative damage to nucleic acids. Markers of both RNA and DNA damage were significantly increased in the smokers with and without CKD compared to their respective control subjects. These findings suggest that a highly specific and sensitive analytical method such as isotope dilution HPLC-MS/MS may represent a valuable tool for the measurement of oxidative stress in human subjects.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Carbon Isotopes; Chromatography, High Pressure Liquid; Deoxyguanosine; DNA Damage; Guanine; Guanosine; Humans; Indicator Dilution Techniques; Kidney Failure, Chronic; Molecular Structure; Nitrogen Isotopes; Oxidation-Reduction; Oxidative Stress; Pyrimidines; Reference Standards; Smoking; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry

Iron, a key element in Fenton chemistry, causes oxygen-related toxicity to cells of most living organisms. Helicobacter pylori is a microaerophilic bacterium that infects human gastric mucosa and causes a series of gastric diseases. Exposure of H. pylori cells to air for 2 h elevated the level of free iron by about 4-fold as measured by electron paramagnetic resonance spectroscopy. H. pylori cells accumulated more free iron as they approached stationary phase growth, and they concomitantly suffered more DNA damage as indicated by DNA fragmentation analysis. Relationships between the intracellular free iron level, specific oxidative stress enzymes, and DNA damage were identified, and new roles for three oxidative stress-combating enzymes in H. pylori are proposed. Mutant cells defective in either catalase (KatA), in superoxide dismutase (SodB) or in alkyl hydroperoxide reductase (AhpC) were more sensitive to oxidative stress conditions; and they accumulated more free (toxic) iron; and they suffered more DNA fragmentation compared to wild type cells. A significant proportion of cells of sodB, ahpC, or katA mutant strains developed into the stress-induced coccoid form or lysed; they also contained significantly higher amounts of 8-oxo-guanine associated with their DNA, compared to wild type cells.

Topics: Air; Bacterial Proteins; Catalase; Cells, Cultured; DNA Damage; DNA Fragmentation; Electron Spin Resonance Spectroscopy; Electrophoresis; Fluorescent Dyes; Free Radicals; Gastric Mucosa; Guanine; Guanosine; Helicobacter pylori; Humans; Iron; Magnetics; Microscopy, Fluorescence; Models, Chemical; Mutation; Oxidative Stress; Oxygen; Peroxidases; Peroxiredoxins; Superoxide Dismutase; Time Factors

8-hydroxy-2'-deoxyguanosine (8OHdG) is a widely used biomarker for the measurement of endogenous oxidative DNA damage. A sensitive method for the quantification of 8OHdG in urine by single solid-phase extraction and GC-MS (gas chromatography with MS detection) using selective ion monitoring is described in the present study. After solid-phase extraction, samples are freeze-dried, derivatized by trimethylsilylation and analysed by GC-MS. The urinary 8OHdG was quantified using heavy isotope dilution with [18O]8OHdG. The recovery of 8OHdG after the solid-phase extraction ranged from 70 to 80% for a wide range of urinary 8OHdG levels. Using 1 ml of urine, the limit of quantification was >2.5 nM (2.5 pmol/ml) and the calibration curve was linear in the range 2.5-200 nM. This method was applied to measure 8OHdG in urine samples from 12 healthy subjects. The intra- and inter-day variations were <9%. Urinary 8OHdG levels in spot urine samples from four healthy subjects were also measured for 1 week and, again, the variation was small. The presence of H2O2 in urine did not cause artifactual formation of 8OHdG. Since this assay is simple, rapid, sensitive and reproducible, it seems suitable to be used as a routine methodology for the measurement of urinary excretion of 8OHdG in large population studies.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Biomarkers; Creatinine; Deoxyguanosine; DNA Damage; Female; Gas Chromatography-Mass Spectrometry; Guanine; Guanosine; Humans; Hydrogen Peroxide; Male; Oxidative Stress; Reproducibility of Results; Spectrometry, Mass, Electrospray Ionization; Time Factors

Crocidolite asbestos is associated with the development of mesothelioma. Although chromosomal changes have been documented in mesothelial cells, the mechanisms of interaction of crocidolite with DNA remain obscure. Since human mesothelial cells are exquisitely sensitive to asbestos, oxidative DNA damage was measured in an asbestos-exposed human mesothelial cell line (MET5A) by assaying oxidized guanine bases [8-oxo-2'-deoxyguanosine (oxo8dG), 8-oxoguanine (oxo8G), and 8-oxoguanosine (oxo8Gua)] excreted into the spent culture medium after DNA repair or turnover. At growth inhibitory, but not cytolytic concentrations, asbestos caused significant elevation of all bases in the spent medium over a 48-h period. In contrast, riebeckite, a chemically similar, nonfibrous analog of crocidolite did not cause increased adduct release. Results show that oxidative RNA and DNA bases are produced in response to asbestos in target cells of asbestos-induced cancers.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Asbestos, Crocidolite; Biomarkers; Carcinogens; Cell Line; Culture Media; Deoxyguanosine; DNA Damage; Epithelium; Guanine; Guanosine; Humans; Oxidation-Reduction; Pleura

Topics: 8-Hydroxy-2'-Deoxyguanosine; Chromatography, Gel; Chromatography, High Pressure Liquid; Deoxyguanosine; Guanine; Guanosine; Indicators and Reagents; Light; Methylene Blue; Photochemistry