Oxidative Stress Levels and DNA Repair Kinetics in Senescent Primary Human Fibroblasts Exposed to Chronic Low Dose Rate of Ionizing Radiation.

BACKGROUND: Exposure to low dose rate (LDR) radiation may accelerate aging processes. Previously, we identified numerous LDR-induced pathways involved in oxidative stress (OS) and antioxidant systems, suggesting that these pathways protect against premature senescence (PS). This study aimed to investigate if there are differences between young replicative senescent (RS) and PS cells considering DNA repair kinetics, OS, and DNA damage localized in the telomeres. METHODS: We established PS cells by culturing and passaging young primary fibroblasts exposed to LDR. Then, RS cells were established by culturing and passaging young fibroblasts until they stopped proliferating. Senescence was characterized by analyzing telomere length and senescence-associated ß-galactosidase (SA-ß-gal) staining. DNA damage and repair were evaluated with γH2AX foci formation; telomere identification was carried out using the fluorescence in situ hybridization (FISH) probe; and oxidative stress was assessed by measuring 8-oxo-dG in the medium. RESULTS: The data indicate the following: young cells have a better ability to cope with LDR-induced oxidative stress; RS and PS have higher steady-state levels of DNA damage; RS have slower DNA repair kinetics; and PS/RS have elevated levels of telomeric DNA damage. CONCLUSION: Our main conclusion is that PS and RS differ regarding DNA repair kinetics and SA-ß-gal levels.

The Influence of 5',8-Cyclo-2'-Deoxyguanosine on ds-DNA Charge Transfer Depends on Its Diastereomeric Form: A Theoretical Study.

The genetic information stored in the nucleobase sequence is continuously exposed to harmful extra- and intra-cellular factors, which can lead to different types of DNA damage, with more than 70 lesion types identified so far. In this article, the influence of a multi-damage site containing (5'R/S) 5',8-cyclo-2'-deoxyguanosine (cdG) and 7,8-dihydro-8-oxo-2'-deoxyguanosine (OXOdG) on charge transfer through ds-DNA was taken into consideration. The spatial geometries of oligo-RcdG: d[A1(5'R)cG2A3OXOG4A5]*d[T5C4T3C2T1] and oligo-ScdG: d[A1(5'S)cG2A3OXOG4A5]*d[T5C4T3C2T1] were optimized at the M06-2X/6-D95**//M06-2X/sto-3G level of theory in the aqueous phase using ONIOM methodology. For all the electronic property energies under discussion, the M06-2X/6-31++G** level of theory was used. Additionally, the non-equilibrated and equilibrated solvent-solute interactions were into consideration. The obtained results confirm the predisposition of OXOdG to radical cation formation regardless of the presence of other lesions in a ds-DNA structure. In the case of electron transfer, however, the situation is different. An excess electron migration towards (5'S)cdG was found to be preferred in the case of oligo-ScdG, while in the case of oligo-RcdG, OXOdG was favored. The above observation was confirmed by the charge transfer rate constant, vertical/adiabatic ionization potential, and electron affinity energy values, as well as the charge and spin distribution analysis. The obtained results indicate that 5',8-cyclo-2'-deoxyguanosine, depending on the C5' atom chirality, can significantly influence the charge migration process through the double helix. The above can be manifested by the slowdown of DNA lesion recognition and removal processes, which can increase the probability of mutagenesis and subsequent pathological processes. With regard to anticancer therapy (radio/chemo), the presence of (5'S)cdG in the structure of formed clustered DNA damage can lead to improvements in cancer treatment.

Telomere fragility in radiology workers occupationally exposed to low doses of ionising radiation.

Ionising radiation damages DNA directly and indirectly through increased production of reactive oxygen species. Although telomeres have been reported as indicators of radiosensitivity, their maintenance in response to occupational exposure to low radiation doses is still a matter of debate. In this work we aimed to investigate telomere length and structure in hospital workers occupationally exposed to X-rays and to relate these findings to oxidation of biomolecules and chromosome aberrations. Blood samples of exposed participants and matching controls were taken during periodical check-ups. Chromosome aberrations and telomere length and structure were analysed in peripheral blood lymphocytes using Q-FISH, whereas oxidative stress parameters [pro/antioxidant balance (PAB), lipid peroxidation, and 8-oxo-dG] were measured in plasma samples. Based on the CA findings we divided the exposed group into two subgroups, of which one had chromosome aberrations in the first division metaphases and the other did not. There was no significant difference in telomere length between any of the groups. However, both subgroups showed significantly higher rate of fragile telomeres and higher lipid peroxidation product and 8-oxo-dG levels than controls. The rate of fragile telomeres significantly correlated with plasma levels of 8-oxo-dG, which suggests that continuous exposure to low radiation doses induces oxidative base damage of guanine resulting in telomere fragility.

Arsenic exposure and biomarkers for oxidative stress and telomere length in indigenous populations in Bolivia.

BACKGROUND: Women living in the Bolivian Andes are environmentally exposed to arsenic, yet there is scarce information about arsenic-related effects in this region. Several biomarkers for telomere length and oxidative stress (mitochondrial DNA copy number, mtDNAcn; 8-Oxo-2'-deoxyguanosine, 8-oxo-dG; and 4-hydroxy nonenal mercapturic acid, 4-HNE-MA) have been previously linked to arsenic, and some of which are prospective biomarkers for cancer risk. OBJECTIVE AND HYPOTHESIS: To evaluate associations between arsenic exposure and telomere length, mtDNAcn, 8-oxo-dG, and 4-HNE-MA in Bolivians. Arsenic exposure was hypothesized to be positively associated with all four toxicity biomarkers, particularly in individuals with a less efficient arsenic metabolism. METHODS: The study encompassed 193 indigenous women. Arsenic exposure was assessed in urine as the sum of inorganic arsenic metabolite concentrations (U-As) measured by HPLC-HG-ICP-MS, and in whole blood as total arsenic (B-As) measured by ICP-MS. Efficiency of arsenic metabolism was evaluated by a polymorphism (rs3740393) in the main arsenic methylating gene AS3MT measured by TaqMan allelic discrimination, and by the relative fractions of urinary inorganic arsenic metabolites. Telomere length and mtDNAcn were determined in peripheral blood leukocytes by quantitative PCR, and urinary 8-oxo-dG and 4-HNE-MA by LC-MS/MS. RESULTS: U-As and B-As were associated with longer telomeres and higher mtDNAcn, particularly in women with a less efficient arsenic metabolism. Urinary 8-oxo-dG and 4-HNE-MA were positively associated with U-As, but only 4-HNE-MA was associated with B-As. Arsenic metabolism efficiency did not have a clear effect on the concentrations of either of these biomarkers. CONCLUSION: Bolivian women showed indications of arsenic toxicity, measured by four different biomarkers. Telomere length, mtDNAcn, and 4-HNE-MA were positively associated with both U-As and B-As. The association of arsenic exposure with telomere length and mtDNAcn was only present in Bolivian women with a less efficient metabolism. These findings call for additional efforts to evaluate and reduce arsenic exposure in Bolivia.

Extra Virgin Olive Oil consumption from Mild Cognitive Impairment patients attenuates oxidative and nitrative stress reflecting on the reduction of the PARP levels and DNA damage.

Oxidative/nitrative stress that results from the unbalance of the overproduction/clearance of reactive oxygen/nitrogen species (ROS/NOS), originated from a variety of endo- and/or exo-genous sources, can have detrimental effects on DNA and is involved in Alzheimer's disease (AD) pathology. An excellent marker of oxidative DNA lesions is 8-hydroxy-2'-deoxyguanosine (8-OHdG) while of nitrative stress the enzyme NOS2 (Nitric oxide synthase 2). Under massive oxidative stress, poly(ADP-ribose)polymerase 1 (PARP-1) enzyme activity, responsible for restoration of DNA damage, is augmented, DNA repair enzymes are recruited, and cell survival/or death is ensued through PARP-1 activation, which is correlated positively with neurodegenerative diseases. In this biochemical study the levels of PARP-1, 8-oxo-dG, and NOS2, Aß1-42, and p-tau in their sera determined using Enzyme-Linked Immunosorbent Assay (ELISA). Patients diagnosed with Mild Cognitive Impairment participated in MICOIL clinical trial, were daily administered with 50 ml Extra Virgin Olive Oil (EVOO) for one year. All MCI patients' biomarkers that had consumed EVOO were tantamount to those of healthy participants, contrary to MCI patients who were not administered. EVOO administration in MCI patients resulted in the restoration of DNA damage and of the well-established "hallmarks" AD biomarkers, thanks probably to its antioxidant properties exhibiting a therapeutic potentiality against AD. Molecular docking simulations of the EVOO constituents on the crystal structure of PARP-1 and NOS-2 target enzymes were also employed, to study in silico the ability of the compounds to bind to these enzymes and explain the observed in vitro activity. In silico analysis has proved the binding of EVOO constituents on PARP-1and NOS-2 enzymes and their interaction with crucial amino acids of the active sites. CLINICAL TRIAL REGISTRATION: https://clinicaltrials.gov/ct2/show/NCT03362996. MICOIL GOV IDENTIFIER: NCT03362996.

Cobalt-induced oxidative stress contributes to alveolar/bronchiolar carcinogenesis in B6C3F1/N mice.

Rodent alveolar/bronchiolar carcinomas (ABC) that arise either spontaneously or due to chemical exposure are similar to a subtype of lung adenocarcinomas in humans. B6C3F1/N mice and F344/NTac rats exposed to cobalt metal dust (CMD) by inhalation developed ABCs in a dose dependent manner. In CMD-exposed mice, the incidence of Kras mutations in ABCs was 67% with 80% of those being G to T transversions on codon 12 suggesting a role of oxidative stress in the pathogenesis. In vitro studies, such as DMPO (5,5-dimethyl-1-pyrroline N-oxide) immune-spin trapping assay, and dihydroethidium (DHE) fluorescence assay on A549 and BEAS-2B cells demonstrated increased oxidative stress due to cobalt exposure. In addition, significantly increased 8-oxo-dG adducts were demonstrated by immunohistochemistry in lungs from mice exposed to CMD for 90 days. Furthermore, transcriptomic analysis on ABCs arising spontaneously or due to chronic CMD-exposure demonstrated significant alterations in canonical pathways related to MAPK signaling (IL-8, ErbB, Integrin, and PAK pathway) and oxidative stress (PI3K/AKT and Melatonin pathway) in ABCs from CMD-exposed mice. Oxidative stress can stimulate PI3K/AKT and MAPK signaling pathways. Nox4 was significantly upregulated only in CMD-exposed ABCs and NOX4 activation of PI3K/AKT can lead to increased ROS levels in human cancer cells. The gene encoding Ereg was markedly up-regulated in CMD-exposed mice. Oncogenic KRAS mutations have been shown to induce EREG overexpression. Collectively, all these data suggest that oxidative stress plays a significant role in CMD-induced pulmonary carcinogenesis in rodents and these findings may also be relevant in the context of human lung cancers.

Antitumour activity of TH1579, a novel MTH1 inhibitor, against castration-resistant prostate cancer.

Castration-resistant prostate cancer (CRPC) treatment still remains difficult. The aim of the present study was to determine the antitumour efficacy of the MutT homolog 1 (MTH1) inhibitor, TH1579, against castration-resistant prostate cancer. PC-3 and DU-145 prostate cancer cells were treated with different concentrations of TH1579. C4-2 cells with or without androgen receptor (AR) were also treated with TH1579 to assess AR function. Cell survival, 8-oxo-dG levels and DNA damage were measured using cell viability assays, western blotting, immunofluorescence analysis and flow cytometry. TH1579 inhibited CRPC cell proliferation in a dose-dependent manner. The viabilities of PC-3 and DU-145 cells treated with 1 µM of TH1579 were 28.6 and 24.1%, respectively. The viabilities of C4-2 cells with and without AR treated with 1 µM TH1579 were 10.6 and 19.0%, respectively. Moreover, TH1579 treatment increased 8-oxo-dG levels, as well as the number of 53BP1 and γH2A.X foci, resulting in increased DNA double-strand breakage and apoptosis in PC-3 and DU-145 cells. The findings of the present study demonstrated that TH1579 exerted strong antitumour effects on CRPC cells, and may therefore be used as a potential therapeutic agent for the clinical treatment of CRPC.

Alterations in levels of 8-Oxo-2'-deoxyguanosine and 8-Oxoguanine DNA glycosylase 1 during a current episode and after remission in unipolar and bipolar depression.

INTRODUCTION: Previous studies showed significant increases in DNA base damage markers and significant alterations in base excision repair enzymes in patients with unipolar and bipolar depression. We aimed to investigate changes in urine 8-Oxo-2'-deoxyguanosine (8-oxo-dG) and gene expression levels of 8-Oxoguanine DNA glycosylase 1 (OGG1) during a current depressive episode and after remission in bipolar and unipolar disorders. METHODS: Twenty-four acutely depressed bipolar (BD), 33 unipolar depression (UD) patients and 61 healthy controls were included in the study. Clinical evaluations, blood and urine sampling were completed at baseline and at remission after eight weeks. The urine 8-oxo-dG levels were assessed by liquid chromatography tandem mass spectrometry and adjusted for urine creatinine levels. The gene expression levels of OGG1 were determined from cDNA extracted from blood samples, using real time-polymerase chain reaction. RESULTS: At baseline, patients presented significantly higher levels of 8-oxo-dG (p = 0.008), and lower gene expression of OGG1 (p = 0.024) compared to controls. Levels of either 8-oxo-dG or OGG1 expression did not differ between BD and UD. In patients who remitted by the 8th week (n = 30), 8-oxo-dG decreased significantly (p = 0.001), and gene expression levels of OGG1 increased by 2.95 times compared to baseline levels (p = 0.001). All comparisons were adjusted for age, sex, smoking status and body mass index. CONCLUSION: Our results suggest that patients with bipolar and unipolar mood disorders present increased 8-oxo-dG and decreased gene expression levels of OGG1 in current depressive episodes, and that these changes might be reversed by the resolution of depressive symptoms. The causal relationship between DNA damage and repair requires further exploration.

Differential Expression Profiles of Oxidative Stress Levels, 8-oxo-dG and 4-HNE, in Barrett's Esophagus Compared to Esophageal Adenocarcinoma.

Barrett's esophagus (BE), a chronic inflammatory condition, is the leading risk factor for esophageal adenocarcinoma (EAC). In inflammation to cancer pathways, oxidative stress profiles have been linked to cancer progression. However, the relevance of oxidative stress profiles along the BE-disease sequence remains to be elucidated. In this study, markers of oxidative stress; DNA adducts (8-oxo-dG) and lipoperoxidation (4-HNE), and markers of proliferation (Ki67) were measured in patient biopsies representing the BE-disease sequence. Differences in expression of these markers in Barrett's patients with cancer-progression and non-progression were examined. Proliferation was reduced in Barrett's specialized intestinal metaplasia (SIM) compared with EAC (p < 0.035). Correcting for cell proliferation levels, a confounding factor, linked to oxidative stress profiles, SIM demonstrated increased levels of 8-oxo-dG and 4-HNE (p < 0.05) compared with EAC. Longitudinal analysis of Barrett's patients demonstrated decreased levels of 8-oxo-dG in SIM cancer progression (p < 0.05). BE is an environment of increased oxidative stress and inflammation. Patients with progressive disease demonstrated reduced oxidative stress levels in 8-oxo-dG. Perhaps these alterations facilitate Barrett's progression, whereas in non-progressive disease, cells follow the rules of increased oxidative stress ultimately triggers cell apoptosis, thereby preventing propagation and survival.

Oxidative Stress: Role and Response of Short Guanine Tracts at Genomic Locations.

Over the decades, oxidative stress has emerged as a major concern to biological researchers. It is involved in the pathogenesis of various lifestyle-related diseases such as hypertension, diabetes, atherosclerosis, and neurodegenerative diseases. The connection between oxidative stress and telomere shortening via oxidative guanine lesion is well documented. Telomeres are confined to guanine rich ends of chromosomes. Owing to its self-association properties, it adopts G-quadruplex structures and hampers the overexpression of telomerase in the cancer cells. Guanine, being the most oxidation prone nucleobase, when structured in G-quadruplex entity, is found to respond peculiarly towards oxidative stress. Interestingly, this non-Watson-Crick structural feature exists abundantly in promoters of various oncogenes, exons and other genomic locations. The involvement of G-quadruplex architecture in oncogene promoters is well recognized in gene regulation processes. Development of small molecules aimed to target G-quadruplex structures, have found to alter the overexpression of oncogenes. The interaction may lead to the obstruction of diseased cell having elevated level of reactive oxygen species (ROS). Thus, presence of short guanine tracts (Gn) forming G-quadruplexes suggests its critical role in oxidative genome damage. Present review is a modest attempt to gain insight on the association of oxidative stress and G-quadruplexes, in various biological processes.

The Effect of Moderate- Versus High-Intensity Resistance Training on Systemic Redox State and DNA Damage in Healthy Older Women.

This study investigated effects of a 16-week progressive resistance training program (RTP) with elastic bands at two different intensities on systemic redox state, DNA damage, and physical function in healthy older women. METHODS: Participants were randomly assigned to the high-intensity group (HIGH; n = 39), moderate-intensity group (MOD; n = 31), or control group (CG; n = 23). The exercise groups performed an RTP twice a week with three to four sets of 6 (HIGH) or 15 (MOD) repetitions of six overall body exercises at a perceived exertion rate of 8-9 on the OMNI-Resistance Exercise Scale for use with elastic bands. Thiol redox state was determined by reduced glutathione (GSH), oxidized glutathione (GSSG), and GSSG/GSH in blood mononuclear cells. Degree of DNA damage was assessed by presence of the oxidized DNA base molecule 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-OHdG) in urine. Physical function monitoring was based on the arm curl, chair stand, up and go, and 6-min walk tests. RESULTS: The HIGH group showed a significant increase in 8-OHdG (+71.07%, effect size [ES] = 1.12) and a significant decrease in GSH (-10.91, ES = -0.69), while the MOD group showed a significant decrease in 8-OHdG levels (-25.66%, ES = -0.69) with no changes in thiol redox state. GSH levels differed significantly between the HIGH and CG groups posttest. The exercise groups showed significant improvements in physical function with no differences between groups. CONCLUSION: RTP at a moderate rather than high intensity may be a better strategy to reduce DNA damage in healthy older women while also increasing independence.

Oxidative imbalance in low/intermediate-1-risk myelodysplastic syndrome patients: The influence of iron overload.

OBJECTIVE: To assess the generation of reactive oxygen species (ROS) and the involvement of the main antioxidant pathways in low/intermediate-1-risk myelodysplastic syndromes (MDS) with iron overload (IOL). METHODS: We examined the levels of superoxide anion (O2-), hydrogen peroxide (H2O2), antioxidants (glutathione, GSH; superoxide dismutase, SOD; catalase, CAT; and glutathione peroxidase, GPx), mitochondrial membrane potential (ΔΨm), and by-products of oxidative damage (8-isoprostanes and 8-oxo-7,8-dihydro-2'-deoxyguanosine, 8-oxo-dG) in 42 MDS patients (28 without IOL at diagnosis, and 14 who developed IOL) and 20 healthy subjects. RESULTS: Patients with IOL showed higher O2- levels (39.4 MFI) than normal controls (22.7 MFI, p=0.0356) and patients at diagnosis (19.4 MFI, p=0.0049). Antioxidant systems, except SOD activity, exhibited significant changes in IOL patients with respect to controls (CAT: 7.1 vs 2.7nmol/ml/min, p=0.0023; GPx: 50.9 vs 76.4nmol/ml/min, p=0.0291; GSH: 50.2 vs 24.1 MFI, p=0.0060). Furthermore, mitochondrial dysfunction was only detected in IOL cases compared to controls (ΔΨm: 3.6 vs 6.4 MFI, p=0.0225). Finally, increased levels of 8-oxo-dG were detected in both groups of patients. CONCLUSION: Oxidative stress is an important but non-static phenomenon in MDS disease, whose status is influenced by, among other factors, the presence of injurious iron.

Influence of genetic background and oxidative stress response on risk of mandibular osteoradionecrosis after radiotherapy of head and neck cancer.

BACKGROUND: Osteoradionecrosis (ORN) of the mandible is a severe complication of head and neck radiotherapy (RT) treatment, where the impact of individual radiosensitivity has been a suggested explanation. METHODS: A cohort of patients with stage II/III ORN was compared to matched controls. Blood was collected and irradiated in vitro to study the capacity to handle radiation-induced oxidative stress. Patients were also genotyped for 8 single-nucleotide polymorphisms (SNPs) in genes involved in the oxidative stress response. RESULTS: A difference in 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) levels was found between the patient cohorts (p = 0.01). The SNP rs1695 in glutathione s-transferase p1 (GSTP1) was also found to be more frequent in the patients with ORN (p = .02). Multivariate analysis of the clinical and biological factors revealed concomitant brachytherapy plus the 2 biomarkers to be significant factors which influense risk of mandibular osteoradionecrosis after radiotherapy of head and neck cancer. CONCLUSION: The current study indicates that oxidative stress response contributes to individual radiosensitivity and healthy tissue damage caused by RT and may be predicted by biomarker analysis.

DNA oxidation profiles of copper phenanthrene chemical nucleases.

The deleterious effects of metal-catalyzed reactive oxygen species (ROS) in biological systems can be seen in a wide variety of pathological conditions including cancer, cardiovascular disease, aging, and neurodegenerative disorder. On the other hand however, targeted ROS production in the vicinity of nucleic acids-as demonstrated by metal-activated bleomycin-has paved the way for ROS-active chemotherapeutic drug development. Herein we report mechanistic investigations into the oxidative nuclease activity and redox properties of copper(II) developmental therapeutics [Cu(DPQ)(phen)](2+) (Cu-DPQ-Phen), [Cu(DPPZ)(phen)](2+) (Cu-DPPZ-Phen), and [{Cu(phen)2}2(µ-terph)](terph) (Cu-Terph), with results being compared directly to Sigman's reagent [Cu(phen)2](2+) throughout (phen = 1,10-phenanthroline; DPQ = dipyridoquinoxaline; DPPZ = dipyridophenazine; Terph = terephthalate). Oxidative DNA damage was identified at the minor groove through use of surface bound recognition elements of methyl green, netropsin, and [Co(NH3)6]Cl3 that functioned to control complex accessibility at selected regions. ROS-specific scavengers and stabilizers were employed to identify the cleavage process, the results of which infer hydrogen peroxide produced metal-hydroxo or free hydroxyl radicals ((•)OH) as the predominant species. The extent of DNA damage owing to these radicals was then quantified through 8-oxo-2'-deoxyguanosine (8-oxo-dG) lesion detection under ELISA protocol with the overall trend following Cu-DPQ-Phen > Cu-Terph > Cu-Phen > Cu-DPPZ. Finally, the effects of oxidative damage on DNA replication processes were investigated using the polymerase chain reaction (PCR) where amplification of 120 base pair DNA sequences of varying base content were inhibited-particularly along A-T rich chains-through oxidative damage of template strands.

Radiation-induced formation of purine lesions in single and double stranded DNA: revised quantification.

The formation of oxidative lesions arising from double stranded DNA damage is of major significance to chemical biology from the perspective of application to human health. The quantification of purine lesions arising from γ-radiation-induced hydroxyl radicals (HO(•)) has been the subject of numerous studies, with discrepancies on the measured 5',8-cyclo-2'-deoxyadenosine (cdA) and 5',8-cyclo-2'-deoxyguanosine (cdG) lesions reported by different groups. Here we applied an ameliorative protocol for the analysis of DNA damage with quantitative determination of these lesions via isotope dilution liquid chromatography coupled with tandem mass spectrometry. Tandem-type purine lesions were quantified along with 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dG) and 7,8-dihydro-8-oxo-2'-deoxyadenosine (8-oxo-dA) in single and double stranded DNA, generated during DNA exposure to diffusible HO(•) radicals in the absence or presence of physiological levels of oxygen. The cdA and cdG lesions in absence of oxygen were found ~2 times higher in single than double stranded DNA, with 5'R being ~6.5 and ~1.5 times more predominant than 5'S in cdG and cdA, respectively. Interestingly, in the presence of 5% molecular oxygen the R/S ratios are retained with substantially decreased yields for cdA and cdG, whereas 8-oxo-dA and 8-oxo-dG remain nearly constant. The overall lesion formation follows the order: 8-oxo-dG >> 8-oxo-dA > 5'R-cdG > 5'R-cdA > 5'S-cdA > 5'S-cdG. By this method, there was a conclusive evaluation of radiation-induced DNA purine lesions.

CYP2E1 induction leads to oxidative stress and cytotoxicity in glutathione-depleted cerebellar granule neurons.

Increasing evidence suggests that brain cytochrome P450 (CYP) can contribute to the in situ metabolism of xenobiotics. In the liver, some xenobiotics can be metabolized by CYPs into more reactive products that can damage hepatocytes and induce cell death. In addition, normal CYP activity may produce reactive oxygen species (ROS) that contribute to cell damage through oxidative mechanisms. CYP2E1 is a CYP isoform that can generate ROS leading to cytotoxicity in multiple tissue types. The aim of this study was to determine whether CYP2E1 induction may lead to significant brain cell impairment. Immunological analysis revealed that exposure of primary cerebellar granule neuronal cultures to the CYP inducer isoniazid, increased CYP2E1 expression. In the presence of buthionine sulfoximine, an agent that reduces glutathione levels, isoniazid treatment also resulted in reactive oxygen species (ROS) production, DNA oxidation and cell death. These effects were attenuated by simultaneous exposure to diallyl sulfide, a CYP2E1 inhibitor, or to a mimetic of superoxide dismutase/catalase, (Euka). These results suggest that in cases of reduced antioxidant levels, the induction of brain CYP2E1 could represent a risk of in situ neuronal damage.

Nickel accumulation in lung tissues is associated with increased risk of p53 mutation in lung cancer patients.

Occupational exposure to nickel compounds has been associated with lung cancer. The correlation between high nickel levels and increased risk of lung cancer has been previously reported in a case-control study. This study assessed whether nickel exposure increased the occurrence of p53 mutations due to DNA repair inhibition by nickel. A total of 189 lung cancer patients were enrolled to determine nickel levels in tumor-adjacent normal lung tissues and p53 mutation status in lung tumors through atomic absorption spectrometry and direct sequencing, respectively. Nickel levels in p53 mutant patients were significantly higher than those in p53 wild-type patients. When patients were divided into high- and low-nickel subgroups by median nickel level, the high-nickel subgroup of patients had an odds ratio (OR) of 3.25 for p53 mutation risk relative to the low-nickel subgroup patients. The OR for p53 mutation risk of lifetime non-smokers, particularly females, in the high-nickel subgroup was greater than that in the low-nickel subgroup. To determine whether nickel affected DNA repair capacity, we conducted the host cell reactivation assay in A549 and H1975 lung cancer cells and showed that the DNA repair activity was reduced by nickel chloride in a dose-dependent manner. This was associated with elevated production of hydrogen peroxide-induced 8-oxo-deoxyguanosine. Therefore, increased risk of p53 mutation due to defective DNA repair caused by high nickel levels in lung tissues may be one mechanism by which nickel exposure contributes to lung cancer development, especially in lifetime female non-smokers.

Oxidative stress in bisphosphonate-related osteonecrosis of the jaws.

OBJECTIVES: To analyze whether oxidative stress (OS) changes are present in patients with bisphosphonate-related osteonecrosis of the jaw (BRONJ) versus controls. MATERIALS AND METHODS: Oxidative stress was analyzed in serum and unstimulated saliva of three groups: Group 1 consisted of 24 patients who had been treated with intravenous bisphosphonates (ivBPs) and developed BRONJ, group 2 consisted of 20 patients who had received ivBPs and did not develop BRONJ, and group 3 comprised 17 control subjects. Reduced glutathione (GSH), malondialdehyde (MDA), oxidized glutathione (GSSG), and 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxo-dG) levels, as well as the GSSG/GSH ratio, were measured. RESULTS: Mean serum and saliva levels of MDA, GSSG, and 8-oxo-dG and the GSSG/GSH ratio were significantly higher in patients with BRONJ than in controls. We found no significant difference in OS according to BRONJ stage, sex, or location in the jaws. Logistic regression analysis revealed that the GSSG/GSH ratio was a significant factor predicting the development of BRONJ (P = 0.01). CONCLUSIONS: Oxidative stress was detected in patients with BRONJ, and the GSSG/GSH ratio was the most significant OS variable found; it was a significant factor predicting the development of BRONJ.

Analysis of mutant frequencies and mutation spectra in hMTH1 knockdown TK6 cells exposed to UV radiation.

Ultraviolet radiation is a highly mutagenic agent that damages the DNA by the formation of mutagenic photoproducts at dipyrimidine sites and by oxidative DNA damages via reactive oxygen species (ROS). ROS can also give rise to mutations via oxidation of dNTPs in the nucleotide pool, e.g. 8-oxo-dGTP and 2-OH-dATP and subsequent incorporation during DNA replication. Here we show that expression of human MutT homolog 1 (hMTH1) which sanitizes the nucleotide pool by dephosphorylating oxidized dNTPs, protects against mutagenesis induced by long wave UVA light and by UVB light but not by short wave UVC light. Mutational spectra analyses of UVA-induced mutations at the endogenous Thymidine kinase gene in human lymphoblastoid cells revealed that hMTH1 mainly protects cells from transitions at GC and AT base pairs.

Level of primary DNA damage in the early stage of metabolic syndrome.

Metabolic syndrome (MetS) is a multi-component disease, characterised by abdominal obesity, hypertension, hyperglycaemia and dyslipidaemia. Since the number of MetS patients has significantly increased over the past two decades and because MetS may lead to development of cardiovascular diseases, diabetes type-2, and cancer, it has become important to extend the knowledge on the pathogenesis of MetS and to establish its possible early biomarkers. Studies on MetS and DNA damage are few and are inconclusive. The aim of this study was to elucidate the involvement of DNA damage in the development of MetS and to establish if DNA damage can serve as early biomarker of MetS. A total of 121 subjects participated in the study: 56 healthy controls and 65 MetS patients who were diagnosed with MetS for the first time. The amount of primary DNA damage in peripheral leukocytes of the subjects was assessed with three types of comet assay: the alkaline, the hOGG1-modified, and the neutral comet assay. In addition, the extent of oxidative DNA damage was monitored in urine by assessing 8-oxo-dG. The parameters of the three types of comet assay did not differ between the control and the MetS group. Interestingly, urinary 8-oxo-dG level in the control group was higher than in the MetS group. Our results imply that DNA damage is not involved in the early stage of MetS and, therefore, DNA damage cannot serve as an early marker of MetS.