L-DOPA oxidation products prevent H2O2-induced oxidative damage to cellular DNA

Using the single-cell gel electrophoresis ("Comet") assay, we show that tyrosinase-generated L-DOPA oxidation products prevent H2O2-induced oxidative DNA damage in cultured tissue cells. We propose that these oxidation products trigger cellular processes that up-regulate the overall antioxidant status of the cell, and could be incorporated into treatments of pathological conditions associated with elevated oxidative DNA damage and other manifestations of increased oxidative stress.     

Antioxidant properties of the mucus secreted by Laeonereis acuta (Polychaeta, Nereididae): a defense against environmental pro-oxidants?

Polychaeta species like Laeonereis acuta (Nereididae) usually secrete great amounts of mucus that wrap the animal inside. Taking into account that fungi action in the sediment and UV radiation acting on dissolved organic matter in the water produces reactive oxygen species (ROS) like hydrogen peroxide (H(2)O(2)), it was considered that the mucus secretion could represent an antioxidant defense against environmental ROS. Antioxidant enzymes (catalase-CAT; superoxide dismutase-SOD; glutathione peroxidase-GPx and glutathione-S-transferase-GST) and total antioxidant capacity (TOSC) were determined in worms and mucus secretion. Higher (p<0.05) CAT, GPx and TOSC values were registered in mucus samples respect worms, SOD activity was similar (p>0.05) in both kind of samples, and absence of GST activity was observed in mucus samples, suggesting absence of catalyzed phase II reactions. In assays conducted with hepatoma cell lines exposed to H(2)O(2), it was verified that: (1) mucus co-exposure significantly (p<0.05) lowered DNA damage induced by H(2)O(2); (2) ROS production was significantly (p<0.05) reduced when cells were exposed simultaneously with mucus samples and H(2)O(2) respect H(2)O(2) alone. It can be concluded that the mucus production contributes substantially to the antioxidant defense system of the worm against environmental ROS through the interception or degradation of H(2)O(2), peroxyl and hydroxyl radicals.     

Cytosolic NADP(+)-dependent isocitrate dehydrogenase status modulates oxidative damage to cells

NADPH is an important cofactor in many biosynthesis pathways and the regeneration of reduced glutathione, critically important in cellular defense against oxidative damage. It is mainly produced by glucose 6-phosphate dehydrogenase (G6PD), malic enzyme, and the cytosolic form of NADP(+)-dependent isocitrate dehydrogenase (IDPc). Little information is available about the role of IDPc in antioxidant defense. In this study we investigated the role of IDPc against cytotoxicity induced by oxidative stress by comparing the relative degree of cellular responses in three different NIH3T3 cells with stable transfection with the cDNA for mouse IDPc in sense and antisense orientations, where IDPc activities were 3-4-fold higher and 35% lower, respectively, than that in the parental cells carrying the vector alone. Although the activities of other antioxidant enzymes, such as superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, and G6PD, were comparable in all transformed cells, the ratio of GSSG to total glutathione was significantly higher in the cells expressing the lower level of IDPc. This finding indicates that IDPc is essential for the efficient glutathione recycling. Upon transient exposure to increasing concentrations of H(2)O(2) or menadione, an intracellular source of free radicals and reactive oxygen species, the cells with low levels of IDPc became more sensitive to oxidative damage by H(2)O(2) or menadione. Lipid peroxidation, oxidative DNA damage, and intracellular peroxide generation were higher in the cell-line expressing the lower level of IDPc. However, the cells with the highly over-expressed IDPc exhibited enhanced resistance against oxidative stress, compared to the control cells. This study provides direct evidence correlating the activities of IDPc and the maintenance of the cellular redox state, suggesting that IDPc plays an important role in cellular defense against oxidative stress.     

NF-kappa B mediates the adaptation of human U937 cells to hydrogen peroxide

Low doses of oxidative stress can induce cellular resistance to subsequent higher doses of the same stress. By using human U937 leukemia cells, we previously demonstrated that H(2)O(2) can induce such an adaptive response without elevating the cellular capacity to degrade H(2)O(2), and were able to confer the cells a cross-resistance to an H(2)O(2)-independent lethal stimulus, C(2)-ceramide. In this study, it was found that the adaptation is accompanied by the translocation of cytoplasmic NF-kappa B to the nuclei. This event was promoted or abolished when either IKK alpha or a dominant negative mutant of I kappa B, respectively, was overexpressed. The overexpression of IKK alpha also resulted in the suppression of H(2)O(2)-induced cell death and DNA fragmentation, whereas these events were accelerated by the expression of the I kappa B mutant. The protective effect of IKK alpha was accompanied neither by an elevation of protein levels of various antioxidant enzymes such as catalase, superoxide dismutase, and glutathione peroxidase, nor by an increase in the cellular capacity to consume H(2)O(2). Moreover, the overexpression of IKK alpha resulted in an enhancement of H(2)O(2)-induced resistance to C(2)-ceramide. The overall data suggest that NF-kappa B mediates the H(2)O(2) adaptation induced in a manner independent of H(2)O(2)-degrading activity.     

Role of abscissic acid in water stress-induced antioxidant defense in leaves of maize seedlings

Roles of abscisic acid (ABA) in water stress-induced oxidative stress were investigated in leaves of maize ( Zea mays L.) seedlings exposed to water stress induced by polyethylene glycol (PEG 6000). Treatment with PEG at -0.7 MPa for 12 and 24 h led to a reduction in leaf relative water content (RWC) by 7.8 and 14.1%, respectively. Duration of the osmotic treatments is considered as mild and moderate water stress. The mild water stress caused significant increases in the generation of superoxide radical ( O 2 - ) and hydrogen peroxide (H 2 O 2 ), the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) and the contents of ascorbate (ASC), reduced glutathione (GSH). The moderate water stress failed to further enhance the capacity of antioxidant defense systems, as compared to the mild water stress. The contents of catalytic Fe, which is critical for H 2 O 2 -dependent hydroxyl radical ( •OH) production, and the oxidized forms of ascorbate and glutathione pools, dehydroascorbate (DHA) and oxidized glutathione (GSSG), markedly increased, a significant oxidative damage to lipids and proteins took place under the moderate water stress. Pretreatment with ABA caused an obvious reduction in the content of catalytic Fe and significant increases in the activities of antioxidant enzymes and the contents of non-enzymatic antioxidants, and then significantly reduced the contents of DHA and GSSG and the degrees of oxidative damage in leaves exposed to the moderate water stress. Pretreatment with an ABA biosynthesis inhibitor, tungstate, significantly suppressed the accumulation of ABA induced by water stress, reduced the enhancement in the capacity of antioxidant defense systems, and resulted in an increase in catalytic Fe, DHA and GSSG, and oxidative damage in the water-stressed leaves. These effects were completely prevented by addition of ABA, which raised the internal ABA content. Our data indicate that ABA plays an important role in water stress-induced antioxidant defense against oxidative stress.     

Induction of oxidative stress and DNA damage in cervix in acute treatment with benzo[a]pyrene

Benzo[a]pyrene [B(a)P] is one of the most prevalent environmental carcinogens and genotoxic agents. However, the mechanisms of B(a)P-induced oxidative damage in cervical tissue are still not clear. The present study was to investigate the oxidative stress and DNA damage in cervix of ICR female mice induced by acute treatment with B(a)P. Oxidative stress was assayed by analysis of malondialdehyde (MDA), superoxide anion and H(2)O(2), and antioxidant enzymes. The alkaline single-cell electrophoresis (SCGE) was used to measure DNA damage. The contents of MDA and glutathione (GSH), and the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione S-transferase (GST) were significantly increased in cervix 24, 48 and 72h after B(a)P treatment of a single dose of 12.5 and 25mg/kg, while GSH, CAT, SOD and GST had no significant difference with the dose of 50mg/kg B(a)P at post-treatment time 48 and 72h except for SOD activity at 48h which was significant. The maximum values of SOD, CAT, GST and GSH were peaked at 24h and then decreased gradually while GPx activities and MDA levels persisted for up to 72h. Superoxide anion, H(2)O(2) and DNA damage changed similarly as the activity of SOD, CAT or GST. Additionally, increases of formamidopyrimidine DNA glycosylase (FPG) specific DNA damage were observed and can be greatly rescued by vitamin C pretreatment. Overall, B(a)P demonstrated a time- and dose- related oxidative stress and DNA damage in cervix.     

Role of Abscisic Acid in Water Stress-induced Antioxidant Defense in Leaves of Maize Seedlings

Roles of abscisic acid (ABA) in water stress-induced oxidative stress were investigated in leaves of maize ( Zea mays L.) seedlings exposed to water stress induced by polyethylene glycol (PEG 6000). Treatment with PEG at &#109 0.7 MPa for 12 and 24 h led to a reduction in leaf relative water content (RWC) by 7.8 and 14.1%, respectively. Duration of the osmotic treatments is considered as mild and moderate water stress. The mild water stress caused significant increases in the generation of superoxide radical ( O 2 &#109 ) and hydrogen peroxide (H 2 O 2 ), the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) and the contents of ascorbate (ASC), reduced glutathione (GSH). The moderate water stress failed to further enhance the capacity of antioxidant defense systems, as compared to the mild water stress. The contents of catalytic Fe, which is critical for H 2 O 2 -dependent hydroxyl radical ( &#148 OH) production, and the oxidized forms of ascorbate and glutathione pools, dehydroascorbate (DHA) and oxidized glutathione (GSSG), markedly increased, a significant oxidative damage to lipids and proteins took place under the moderate water stress. Pretreatment with ABA caused an obvious reduction in the content of catalytic Fe and significant increases in the activities of antioxidant enzymes and the contents of non-enzymatic antioxidants, and then significantly reduced the contents of DHA and GSSG and the degrees of oxidative damage in leaves exposed to the moderate water stress. Pretreatment with an ABA biosynthesis inhibitor, tungstate, significantly suppressed the accumulation of ABA induced by water stress, reduced the enhancement in the capacity of antioxidant defense systems, and resulted in an increase in catalytic Fe, DHA and GSSG, and oxidative damage in the water-stressed leaves. These effects were completely prevented by addition of ABA, which raised the internal ABA content. Our data indicate that ABA plays an important role in water stress-induced antioxidant defense against oxidative stress.     

Mechanisms of sulfur mustard analog 2-chloroethyl ethyl sulfide-induced DNA damage in skin epidermal cells and fibroblasts

Employing mouse skin epidermal JB6 cells and dermal fibroblasts, here we examined the mechanisms of DNA damage by 2-chloroethyl ethyl sulfide (CEES), a monofunctional analog of sulfur mustard (SM). CEES exposure caused H2A.X and p53 phosphorylation as well as p53 accumulation in both cell types, starting at 1 h, that was sustained for 24 h, indicating a DNA-damaging effect of CEES, which was also confirmed and quantified by alkaline comet assay. CEES exposure also induced oxidative stress and oxidative DNA damage in both cell types, measured by an increase in mitochondrial and cellular reactive oxygen species and 8-hydroxydeoxyguanosine levels, respectively. In the studies distinguishing between oxidative and direct DNA damage, 1 h pretreatment with glutathione (GSH) or the antioxidant Trolox showed a decrease in CEES-induced oxidative stress and oxidative DNA damage. However, only GSH pretreatment decreased CEES-induced total DNA damage measured by comet assay, H2A.X and p53 phosphorylation, and total p53 levels. This was possibly due to the formation of GSH–CEES conjugates detected by LC-MS analysis. Together, our results show that CEES causes both direct and oxidative DNA damage, suggesting that to rescue SM-caused skin injuries, pleiotropic agents (or cocktails) are needed that could target multiple pathways of mustard skin toxicities.     

Vascular superoxide and hydrogen peroxide production and oxidative stress resistance in two closely related rodent species with disparate longevity

Vascular aging is characterized by increased oxidative stress, impaired nitric oxide (NO) bioavailability and enhanced apoptotic cell death. The oxidative stress hypothesis of aging predicts that vascular cells of long-lived species exhibit lower production of reactive oxygen species (ROS) and/or superior resistance to oxidative stress. We tested this hypothesis using two taxonomically related rodents, the white-footed mouse (Peromyscus leucopus) and the house mouse (Mus musculus), that show a more than twofold difference in maximum lifespan potential (MLSP = 8 and 3.5 years, respectively). We compared interspecies differences in endothelial superoxide (O2-) and hydrogen peroxide (H2O2) production, NAD(P)H oxidase activity, mitochondrial ROS generation, expression of pro- and antioxidant enzymes, NO production, and resistance to oxidative stress-induced apoptosis. In aortas of P. leucopus, NAD(P)H oxidase expression and activity, endothelial and H2O2 production, and ROS generation by mitochondria were less than in mouse vessels. In P. leucopus, there was a more abundant expression of catalase, glutathione peroxidase 1 and hemeoxygenase-1, whereas expression of Cu/Zn-SOD and Mn-SOD was similar in both species. NO production and endothelial nitric oxide synthase expression was greater in P. leucopus. In mouse aortas, treatment with oxidized low-density lipoprotein (oxLDL) elicited substantial oxidative stress, endothelial dysfunction and endothelial apoptosis (assessed by TUNEL assay, DNA fragmentation and caspase 3 activity assays). According to our prediction, vessels of P. leucopus were more resistant to the proapoptotic effects of oxidative stressors (oxLDL and H2O2). Primary fibroblasts from P. leucopus also exhibited less H2O2-induced DNA damage (comet assay) than mouse cells. Thus, increased lifespan potential in P. leucopus is associated with a decreased cellular ROS generation and increased oxidative stress resistance, which accords with the prediction of the oxidative stress hypothesis of aging.     

Protective effects of cyclosativene on H2O2-induced injury in cultured rat primary cerebral cortex cells

Sesquiterpenes have attracted much interest with respect to their protective effect against oxidative damage that may be the cause of many diseases including several neurodegenerative disorders and cancer. Our previous unpublished work suggested that cyclosativene (CSV), a tetracyclic sesquiterpene, has antioxidant and anticarcinogenic features. However, little is known about the effects of CSV on oxidative stress induced neurotoxicity. We used hydrogen peroxide (H₂O₂) exposure for 6 h to model oxidative stress. Therefore, this experimental design allowed us to explore the neuroprotective potential of CSV in H₂O₂-induced toxicity in new-born rat cerebral cortex cell cultures for the first time. For this aim, MTT and lactate dehydrogenase release assays were carried out to evaluate cytotoxicity. Total antioxidant capacity (TAC) and total oxidative stress (TOS) parameters were used to evaluate oxidative changes. In addition to determining of 8-hydroxy-2-deoxyguanosine (8-OH-dG) levels, the single cell gel electrophoresis (or Comet assay) was also performed for measuring the resistance of neuronal DNA to H₂O₂-induced challenge. Our results showed that survival and TAC levels of the cells decreased, while TOS, 8-OH-dG levels and the mean values of the total scores of cells showing DNA damage (Comet assay) increased in the H₂O₂ alone treated cultures. But pre-treatment of CSV suppressed the cytotoxicity, genotoxicity and oxidative stress which were increased by H₂O₂. On the basis of these observations, it is suggested that CSV as a natural product with an antioxidant capacity in mitigating oxidative injuries in the field of neurodegenerative disorders.     

Hyperoside prevents oxidative damage induced by hydrogen peroxide in lung fibroblast cells via an antioxidant effect

We elucidated the cytoprotective effects of hyperoside (quercetin-3-O-galactoside) against hydrogen peroxide (H2O2)-induced cell damage. We found that hyperoside scavenged the intracellular reactive oxygen species (ROS) detected by fluorescence spectrometry, flow cytometry, and confocal microscopy. In addition, we found that hyperoside scavenged the hydroxyl radicals generated by the Fenton reaction (FeSO4)+H2O2) in a cell-free system, which was detected by electron spin resonance (ESR) spectrometry. Hyperoside was found to inhibit H2O2-induced apoptosis in Chinese hamster lung fibroblast (V79-4) cells, as shown by decreased apoptotic nuclear fragmentation, decreased sub-G(1) cell population, and decreased DNA fragmentation. In addition, hyperoside pretreatment inhibited the H2O2-induced activation of caspase-3 measured in terms of levels of cleaved caspase-3. Hyperoside prevented H2O2-induced lipid peroxidation as well as protein carbonyl. In addition, hyperoside prevented the H2O2-induced cellular DNA damage, which was established by comet tail, and phospho histone H2A.X expression. Furthermore, hyperoside increased the catalase and glutathione peroxidase activities. Conversely, the catalase inhibitor abolished the cytoprotective effect of hyperoside from H2O2-induced cell damage. In conclusion, hyperoside was shown to possess cytoprotective properties against oxidative stress by scavenging intracellular ROS and enhancing antioxidant enzyme activity.     

Mitochondrial DNA damage is sensitive to exogenous H(2)O(2) but independent of cellular ROS production in prostate cancer cells

Intrinsic oxidative stress through enhanced production of reactive oxygen species (ROS) in prostate and other cancers may contribute to cancer progression due to its stimulating effect on cancer growth. In this study, we investigate differential responses to exogenous oxidative stimuli between aggressive prostate cancer and normal cell lines and explore potential mechanisms through interactions between cytotoxicity, cellular ROS production and oxidative DNA damage. The circular, multi-copy mitochondrial DNA (mtDNA) is used as a sensitive surrogate to oxidative DNA damage. We demonstrate that exogenous H(2)O(2) induces preferential cytotoxicity in aggressive prostate cancer than normal cells; a cascade production of cellular ROS, composed mainly of superoxide (O(2)(-)), is shown to be a critical determinant of H(2)O(2)-induced selective toxicity in cancer cells. In contrast, mtDNA damage and copy number depletion, as measured by a novel two-phase strategy of the supercoiling-sensitive qPCR method, are very sensitive to exogenous H(2)O(2) exposure in both cancer and normal cell lines. Moreover, we demonstrate for the first time that the sensitive mtDNA damage response to exogenous H(2)O(2) is independent of secondary cellular ROS production triggered by several ROS modulators regardless of cell phenotypes. These new findings suggest different mechanisms underpinning cytotoxicity and DNA damage induced by oxidative stress and a susceptible phenotype to oxidative injury associated with aggressive prostate cancer cells in vitro.     

胶原蛋白肽经由Nrf2信号通路对H_2O_2诱导的肝细胞氧化损伤的保护作用(英文)

目的:氧化应激在肝脏疾病中扮演着重要的角色。胶原蛋白肽是天然的抗氧化剂,其在动物实验中已经被证实有抑制氧化应激的作用。最新研究证实胶原蛋白肽将有可能被应用在肝脏疾病的预防中,但是很少有研究报道其分子作用机制。因此本研究在胶原蛋白肽是对H2O2诱导的正常人的肝细胞系HL7702氧化损伤有保护作用的基础上,并探索其分子作用机制。方法:实验设空白对照组,H2O2模型组,胶原蛋白肽低、中、高剂量组(10,100,200μg/ml)。胶原蛋白肽各组加入相应浓度的药物预处理12 h后,与模型组一起加入300μM H2O2的H2O2共同培养12 h,空白对照组正常培养。细胞毒性是由CCK8和乳酸脱氢酶(LDH)的释放检测。抗氧化试剂盒检测细胞内活性氧的水平,超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性和丙二醛(MDA)含量的变化。Western blot检测细胞内Nrf2蛋白的表达水平。结果:胶原蛋白肽对H2O2诱导的正常人的肝细胞系HL7702氧化损伤有保护作用。胶原蛋白肽能够及时清除细胞内的活性氧,增加Nrf2的蛋白表达水平,提高超氧化物歧化酶(SOD)、过氧化氢酶(CAT)的活性,减轻脂质过氧化反应,从而保护正常人的肝细胞系HL7702。结论:总之,胶原蛋白肽通过增加Nrf2的蛋白表达水平,提高抗氧化活性,对H2O2诱导损伤的肝细胞发挥保护作用。本研究为胶原蛋白肽的分子作用机制提供了新的证据,将有助于预防氧化应激所致的肝损伤。     

Effect of copper deficiency on oxidative DNA damage in Jurkat T-lymphocytes

The micronutrient copper is a catalytic cofactor for copper, zinc superoxide dismutase and ceruloplasmin, which are two important antioxidant enzymes. As such, a lack of copper may promote oxidative stress and damage. The purpose of this study was to determine the effect of copper deficiency on oxidative damage to DNA in Jurkat T-lymphocytes. To induce copper deficiency, cells were incubated for 48 h with 5-20 microM 2,3,2-tetraamine (2,3,2-tet), a high affinity copper chelator. Such treatment did not affect cell proliferation/viability, as assessed by measuring mitochondrial reduction of WST-1 reagent (4-[3-(4-Iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-ben zen e disulfonate). Furthermore, the induction of copper deficiency did not promote oxidative DNA damage as evaluated by the comet assay. Comet scores were 15 +/- 0 and 16 +/- 1 for control and copper-deficient cells, respectively. However, the copper-deficient cells sustained greater oxidative DNA damage than the control cells (comet scores of 175 +/- 15 and 50 +/- 10, respectively) when both were oxidatively challenged with 50 microM hydrogen peroxide (H(2)O(2)). Supplemental copper but not zinc or iron prevented the potentiation of the H(2)O(2)-induced oxidative DNA damage caused by 2,3,2-tet. These data suggest that copper deficiency compromises the antioxidant defense system of cells, thereby increasing their susceptibility to oxidative DNA damage.     

Oxidative damage to proteins in yeast cells exposed to adaptive levels of H(2)O(2)

When yeast cells are exposed to sublethal concentrations of oxidants, they adapt to tolerate subsequent lethal treatments. Here, we show that this adaptation involves tolerance of oxidative damage, rather than protection of cellular constituents. o- and m-tyrosine levels are used as a sensitive measure of protein oxidative damage and we show that such damage accumulates in yeast cells exposed to H(2)O(2) at low adaptive levels. Glutathione represents one of the main cellular protections against free radical attack and has a role in adaptation to oxidative stress. Yeast mutants defective in glutathione metabolism are shown to accumulate significant levels of o- and m-tyrosine during normal aerobic growth conditions.     

Protective effect of butin against hydrogen peroxide-induced apoptosis by scavenging reactive oxygen species and activating antioxidant enzymes

The antioxidant property of butin was investigated for cytoprotective effect against H(2)O(2)-induced cell damage. This compound showed intracellular reactive oxygen species (ROS) scavenging, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, inhibition of lipid peroxidation, and DNA damage. This radical scavenging activity of butin protected cell damage exposed to H(2)O(2). Also, butin reduced the apoptotic cells induced by H(2)O(2), as demonstrated by the decreased DNA fragmentation, apoptotic body formation, and caspase 3 activity. In addition, butin restored the activity and protein expression of cellular antioxidant enzymes, superoxide dismutase (SOD), and catalase (CAT) in H(2)O(2)-treated cells. Taken together, these findings suggest that butin protected cells against H(2)O(2)-induced cell damage via antioxidant property.     

Antioxidant effect of zinc and zinc-metallothionein in the acute cytotoxicity of hydrogen peroxide in Ehrlich ascites tumour cells

This study was concerned with the role of zinc (Zn) and zinc-metallothionein (Zn-MT) in oxidative stress. Hydrogen peroxide-induced oxidative injury was examined in Ehrlich ascites tumour cells isolated from control host mice, mice pretreated with 10 mg/kg ZnSO4 (i.p.) to increase cellular Zn/Zn-MT levels, and mice exposed to Zn-deficient diet to reduce the cellular Zn/Zn-MT levels. The results of the present study showed that Ehrlich cells with seven-fold differences in Zn-MT concentrations could be obtained by manipulating the Zn status of host mice and that high Zn and Zn-MT levels can make Ehrlich cells more resistant to H2O2-induced oxidative injury (cell viability, lipid peroxidation, [Ca2+]i) while cells with reduced Zn/Zn-MT levels were more susceptible to this treatment. H2O2 treatment resulted in oxidation of MT thiolate groups and loss of its metal binding capacity with translocation of Zn released from oxidized MT to other cellular sites. Preincubation of Ehrlich cells with ZnSO4 in vitro also conferred some degree of resistance to H2O2 toxicity, suggesting the inherent antioxidative property of Zn ions. These data suggested that Zn-MT can be considered as an antioxidant by virtue of its thiolate groups and its Zn ions that are released in the presence of oxidative stress.     

Contribution of mitochondrial DNA repair to cell resistance from oxidative stress

Numerous studies have revealed that a part of the cellular response to chronic oxidative stress involves increased antioxidant capacity. However, another defense mechanism that has received less attention is DNA repair. Because of the important homeostatic role of mitochondria and the exquisite sensitivity of mitochondrial DNA (mtDNA) to oxidative damage, we hypothesized that mtDNA repair plays an important role in the protection against oxidative stress. To test this hypothesis mtDNA damage and repair was evaluated in normal HA1 Chinese hamster fibroblasts and oxidative stress-resistant variants isolated following chronic exposure to H2O2 or 95% O2. Reactive oxygen species were generated enzymatically using xanthine oxidase and hypoxanthine. When treated with xanthine oxidase reduced levels of initial mtDNA damage and enhanced mtDNA repair were observed in the cells from the oxidative stress-resistant variants, relative to the parental cell line. This enhanced mtDNA repair correlated with an increase in mitochondrial apurinic/apyrimidinic endonuclease activity in both H2O2- and O2-resistant HA1 variants. This is the first report showing enhanced mtDNA repair in the cellular response to chronic oxidative stress. These results provide further evidence for the crucial role that mtDNA repair pathways play in protecting cells against the deleterious effects of reactive oxygen species.