Ascorbic Acid Inhibits Lipid Peroxidation but Enhances DNA Damage in Rat Liver Nuclei Incubated with Iron Ions

In this report we studied DNA damage and lipid peroxidation in rat liver nuclei incubated with iron ions for up to 2 hrs in order to examine whether nuclear DNA damage was dependent on membrane lipid peroxidation. Lipid peroxidation was measured as thio-barbituric acid-reactive substances (TBARS) and DNA damage was measured as 8-OH-deoxyguanosine (8-OH-dG). We showed that Fe(II) induced nuclear lipid peroxidation dose-dependently but only the highest concentration (1.0 mM) used induced appreciable 8-OH-dG. Fe(II1) up to 1 mM induced minimal lipid peroxidation and negligible amounts of 8-OH-dG. Ascorbic acid enhanced Fe(II)-induced lipid peroxidation at a ratio to Fe(II) of 1:l but strongly inhibited peroxidation at ratios of 2.5:l and 5:l. By contrast, ascorbate markedly enhanced DNA damage at all ratios tested and in a concentration-dependent manner. The nuclear DNA damage induced by 1 niM FeSO₄/5 mM ascorbic acid was largely inhibited by iron chelators and by dimethylsulphoxide and manni-tol, indicating the involvement of OH. Hydrogen peroxide and superoxide anions were also involved, as DNA damage was partially inhibited by catalase and, to a lesser extent, by superoxide dismutase. The chain-breaking antioxidants butylated hydroxytoluene and diphenylamine (an alkoxyl radical scavenger) did not inhibit DNA damage. Hence, this study demonstrated that ascorbic acid enhanced Fe(II)-induced DNA base modification which was not dependent on lipid peroxidation in rat liver nuclei.     

Relation of lipid peroxidation to loss of cations trapped in liposomes

Lipid peroxidation and alterations in cation loss have been induced in liposomes by ferrous ion, ascorbic acid, reduced and oxidized glutathione, and gamma radiation. Modifications of these effects by tocopherol and 2,6-di-tert-butyl-4-methylphenol (BHT) were studied when these antioxidants were either incorporated in the membrane or were added to already formed liposomes prior to the addition of the chemical agent or to irradiation. Lipid peroxidation, as indicated by the thiobarbituric acid test for malonic dialdehyde, did not correlate with alterations in cation loss. The largest amounts of lipid peroxidation induced by ascorbic acid and glutathione were associated with decreased cation loss. Inhibition of Fe(2+)- and radiation-induced lipid peroxidation by antioxidants did not inhibit the associated increase in cation loss. Tocopherol was a more effective antioxidant than BHT when it was incorporated in the membrane, whereas BHT was more effective when it was added to the liposomes after formation.     

DNA damage (comet assay) and 8-oxodGuo (HPLC-EC) in relation to oxidative stress in the freshwater bivalve Unio tumidus.

The relationships between DNA damage and oxidative stress in the digestive gland, gills and haemocytes of the freshwater bivalve Unio tumidus were investigated. Two markers of genotoxicity were measured: DNA breaks by means of the comet assay, and oxidative DNA lesions by means of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) measured using high-performance liquid chromatography (HPLC) coupled to electrochemical detection. Lipid peroxidation was evaluated by measuring malondialdehyde (MDA) tissue levels. Effects were studied after exposure of bivalves for 6 days to benzo[a]pyrene (B[a]P) (50 and 100 microg l(-1)) and ferric iron (20 and 40 mg l(-1)), applied alone or in combination. Lipid peroxidation in the digestive gland and gills resulted from exposure to Fe3+ or B[a]P whatever the concentrations tested. DNA oxidatively formed lesions were induced in the two tissues at a higher level after B[a]P exposure than after Fe3+ treatment. No significant dose-response relationship was found with the two compounds and no synergistic effect was observed between Fe3+ and B[a]P. The gills appeared less sensitive than the digestive gland to DNA lesions expressed as 8-oxodGuo and comet results. Good correlations were noted between 8-oxodGuo and comet. MDA and DNA damage did not correlate as well, although it was stronger in the digestive gland than in the gills. Production of mucus by the gills likely served to prevent lesions by reducing the bioavailability of the chemicals tested, which could explain that dose-effect relationships and synergistic effects were not observed.     

Inflammation, gene mutation and photoimmunosuppression in response to UVR-induced oxidative damage contributes to photocarcinogenesis

Ultraviolet (UV) radiation causes inflammation, gene mutation and immunosuppression in the skin. These biological changes are responsible for photocarcinogenesis. UV radiation in sunlight is divided into two wavebands, UVB and UVA, both of which contribute to these biological changes, and therefore probably to skin cancer in humans and animal models. Oxidative damage caused by UV contributes to inflammation, gene mutation and immunosuppression. This article reviews evidence for the hypothesis that UV oxidative damage to these processes contributes to photocarcinogenesis. UVA makes a larger impact on oxidative stress in the skin than UVB by inducing reactive oxygen and nitrogen species which damage DNA, protein and lipids and which also lead to NAD+ depletion, and therefore energy loss from the cell. Lipid peroxidation induces prostaglandin production that in association with UV-induced nitric oxide production causes inflammation. Inflammation drives benign human solar keratosis (SK) to undergo malignant conversion into squamous cell carcinoma (SCC) probably because the inflammatory cells produce reactive oxygen species, thus increasing oxidative damage to DNA and the immune system. Reactive oxygen or nitrogen appears to cause the increase in mutational burden as SK progress into SCC in humans. UVA is particularly important in causing immunosuppression in both humans and mice, and UV lipid peroxidation induced prostaglandin production and UV activation of nitric oxide synthase is important mediators of this event. Other immunosuppressive events are likely to be initiated by UV oxidative stress. Antioxidants have also been shown to reduce photocarcinogenesis. While most of this evidence comes from studies in mice, there is supporting evidence in humans that UV-induced oxidative damage contributes to inflammation, gene mutation and immunosuppression. Available evidence implicates oxidative damage as an important contributor to sunlight-induced carcinogenesis in humans.     

Wavelength dependence of biological damage induced by UV radiation on bacteria

The biological effects of UV radiation of different wavelengths (UVA, UVB and UVC) were assessed in nine bacterial isolates displaying different UV sensitivities. Biological effects (survival and activity) and molecular markers of oxidative stress [DNA strand breakage (DSB), generation of reactive oxygen species (ROS), oxidative damage to proteins and lipids, and the activity of antioxidant enzymes catalase and superoxide dismutase] were quantified and statistically analyzed in order to identify the major determinants of cell inactivation under the different spectral regions. Survival and activity followed a clear wavelength dependence, being highest under UVA and lowest under UVC. The generation of ROS, as well as protein and lipid oxidation, followed the same pattern. DNA damage (DSB) showed the inverse trend. Multiple stepwise regression analysis revealed that survival under UVA, UVB and UVC wavelengths was best explained by DSB, oxidative damage to lipids, and intracellular ROS levels, respectively.     

Peroxide-induced membrane damage in human erythrocytes

Erythrocytes exposed to H2O2 or t-butyl hydroperoxide (tBHP) exhibited lipid peroxidation and increased passive cation permeability. In the case of tBHP a virtually complete inhibition of both processes was caused by butylated hydroxytoluene (BHT), whereas pretreatment of the cells with CO increased both lipid peroxidation and K+ leakage. In the experiments with H2O2, on the other hand, both BHT and CO strongly inhibited lipid peroxidation, without affecting the increased passive cation permeability. These observations indicate different mechanisms of oxidative damage, induced by H2O2 and tBHP, respectively. The SH-reagent diamide strongly inhibited H2O2-induced K+ leakage, indicating the involvement of SH oxidation in this process. With tBHP, on the contrary, K+ leakage was not significantly influenced by diamide. Thiourea inhibited tBHP-induced K+ leakage, without affecting lipid peroxidation. Together with other experimental evidence this contradicts a rigorous interdependence of tBHP-induced lipid peroxidation and K+ leakage.     

Lipid peroxidation, protein thiol oxidation and DNA damage in hydrogen peroxide-induced injury to endothelial cells: role of activation of poly(ADP-ribose)polymerase

These experiments are a continuation of work investigating the mechanism of oxidant-induced damage to cultured bovine pulmonary artery endothelial cells (BPEC). Earlier experiments implicated DNA strand breakage and activation of poly(ADP-ribose)polymerase as critical steps in cell injury. In the current report, a better defined model of oxidant stress was used to investigate DNA damage, lipid peroxidation and protein thiol oxidation in BPEC following oxidant stress. The dose and time response of LDH release following exposure to H2O2 were established. H2O2 was metabolized rapidly by BPEC (t1/2 = 20 min). Hydrogen peroxide-induced increases in thiobarbituric acid (TBA) reactive material were prevented by pretreatment with the lipophilic antioxidant diphenylphenylinediamine (DPPD). However, DPPD did not decrease LDH release. Conversely, pretreatment with 5 mM 3-aminobenzamide (3AB), a competitive inhibitor of poly(ADP-ribose)polymerase, prevented LDH release from BPEC following H2O2 treatment. Dithiothreitol (DTT), a sulfhydryl reducing agent, also prevented LDH release. The effects of 3AB and DTT on H2O2-induced changes in DNA strand breaks and NAD+ and ATP levels were investigated as well as the effect of H2O2 on soluble and protein-bound thiols. As DPPD inhibited peroxidation without preventing LDH release, lipid peroxidation does not appear to play a role in the loss of BPEC viability in response to oxidant stress. As protein thiol oxidation was not caused by H2O2, it does not appear to play a causative role in cytotoxicity, although DTT may protect via maintenance of soluble thiols. H2O2 induces DNA strand breaks, which activate poly(ADP-ribose)polymerase, leading to depletion of cellular NAD+ and ATP and loss in cell viability. This supports earlier studies implicating the activation of poly(ADP-ribose)polymerase in oxidant injury to cultured endothelial cells.     

Characteristics of Fe(II)ATP complex-induced damage to the rat liver mitochondrial membrane

It is well established that several iron complexes can induce oxidative damage in hepatic mitochondrial membranes by catalyzing the formation of ·OH radicals and/or by promoting lipid peroxidation. This is a relevant process for the molecular basis of iron overload diseases. The present work demonstrates that Fe(II)ATP complexes (5–50M) promote an oxygen consumption burst in a suspension of isolated rat liver mitochondria (either in the absence or presence of Antimycin A), caused mainly by lipid peroxidation. Fe(II)ATP alone induced small levels of oxygen uptake but no burst. The time course of Fe(II)ATP oxidation to Fe(III)ATP in the extramitochondrial media also reveals a simultaneous burst phase. The iron chelator Desferal (DFO) or the chain-break antioxidant butylated hydroxytoluene (BHT) fully prevented both lipid peroxidation (quantified as oxygen uptake burst) and mitochondrial swelling. DFO and BHT were capable of stopping the ongoing process of peroxidation at any point of their addition to the mitochondrial suspension. Conversely, DFO and BHT only halted the Fe(II)ATP-induced mitochondrial swelling at the onset of the process. Fe(II)ATP could also cause the collapse of mitochondrial potential, which was protected by BHT if added at the onset of the damaging process. These results, as well as correlation studies between peroxidation and mitochondrial swelling, suggest that a two phase process is occurring during Fe(II)ATP-induced mitochondrial damage: one dependent and another independent of lipid peroxidation. The involvement of lipid peroxidation in the overall process of mitochondrial membrane injury is discussed.Abbreviations AA Antimycin A - BHT butylated hydroxytoluene - EGTA ethylene glycol-bis(-aminoethyl ether) - N,N,N,N tetraacetic acid - DFO Desferal - HEPES N-(2-hydroxyethyl)piperazine-N-2-ethanesulfonic acid - SOD superoxide dismutase - TPP+ tetraphenylphosphonium bromide - TBARS thiobarbituric acid reactive substances     

Zinc protects against ultraviolet A1-induced DNA damage and apoptosis in cultured human fibroblasts

Ultraviolet Al (UVA1) radiation generates reactive oxygen species and the oxidative stress is known as a mediator of DNA damage and of apoptosis. We exposed cultured human cutaneous fibroblasts to UVA1 radiation (wavelengths in the 340–450-nm range with emission peak at 365 nm) and, using the alkaline unwinding method, we showed an immediate significant increase of DNA strand breaks in exposed cells. Apoptosis was determined by detecting cytoplasmic nucleosomes (enzyme-linked immunosorbent assay method) at different time points in fibroblasts exposed to different irradiation doses. In our conditions, UVA1 radiation induced an early (8 h) and a delayed (18 h) apoptosis. Delayed apoptosis increased in a UVA dosedependent manner. Zinc is an important metal for DNA protection and has been shown to have inhibitory effects on apoptosis. The addition of zinc (6.5 mg/L) as zinc chloride to the culture medium significantly decreased immediate DNA strand breaks in human skin fibroblasts. Moreover, zinc chloride significantly decreased UVA1-induced early and delayed apoptosis. Thus, these data show for the first time in normal cutaneous cultured cells that UVA1 radiation induces apoptosis. This apoptosis is biphasic and appears higher 18 h after the stress. Zinc supplementation can prevent both immediate DNA strand breakage and early and delayed apoptosis, suggesting that this metal could be of interest for skin cell protection against UVA1 irradiation.     

脂质过氧化引起的DNA损伤研究进展

脂质过氧化可以引起各种碱基损伤、DNA链断裂和各种荧光产物生成,并对DNA分子鸟嘌呤碱基具有选择性损伤.过渡金属离子可以明显加深脂质过氧化对DNA的损伤程度.多种抗氧化剂、活性氧自由基清除剂对脂质过氧化引起的DNA损伤有一定程度的保护作用.具有致突、致癌作用的8－羟基鸟嘌呤已经观察到．脂质过氧化的致突变、致癌变作用机制引起了人们的极大兴趣．     

Iron-induced DNA damage and synthesis in isolated rat liver nuclei.

Incubation of iron with isolated rat liver nuclei stimulated fragmentation of single-stranded DNA, incorporation of [3H]thymidine into DNA and the binding of 59Fe to DNA. FeCl2 was about twice as active as FeCl3. Lipid peroxidation took place in nuclei incubated with FeCl2, but not with FeCl3. Generation of reactive forms of oxygen was required for iron-mediated DNA damage, but evidence for direct interaction of reactive oxygen with DNA was not found. Apparent adducts of iron bound to DNA seemed to be formed by an enzymic mechanism.     

DNA damage induced by catechol derivatives

We investigated the effect of catechol derivatives, including dopa, dopamine, adrenaline and noradrenaline, on DNA damage and the mechanisms of DNA strand breakage and formation of 8-hydroxyguanine (8HOG). The catechol derivatives caused strand breakage of plasmid DNA in the presence of ADP-Fe(3+). The DNA damage was prevented by catalase, mannitol and dimethylsulfoxide, suggesting hydroxyl radical (HO..)-like species are involved in the strand breakage of DNA. Iron chelators, such as desferrioxamine and bathophenanthroline, and reduced glutathione also inhibited the DNA damage. Deoxyribose, a molecule that is used to detect HO,, was not degraded by dopa in the presence of ADP-Fe(3+). By adding EDTA, however, dopa induced the marked deoxyribose degradation in the presence of ADP-Fe(3+), indicating that EDTA may extract iron from ADP-Fe(3+) to catalyze HO. formation by dopa. Thus, EDTA was a good catalyst for HO.-generation, whereas it did not promote the strand breakage of DNA. However, calf thymus DNA base damage, which was detected as 8-HOG formation, was caused by dopa in the presence of EDTA-Fe(3+), but not in the presence of ADP-Fe(3+). The 8HOG formation was also inhibited by catalase and HO. scavengers, indicating that HO&z.rad; was involved in the base damage. These results suggest that DNA strand breakage is due to ferryl species rather than HO., and that 8HOG formation is due to HO. rather than ferryl species.     

Vitamin E prevents exercise-induced DNA damage

The single cell gel test (SCG test or comet assay) was used to study DNA damage in peripheral white blood cells (WBC) of humans after a single bout of exhaustive exercise and the effect of vitamin supplementation. Human subjects were asked to run on a treadmill until exhaustion and blood samples were taken before and 24 h after the run. A clear increase in DNA strand breakage was observed in the 24-h sample of all probands. A short-term application of multivitamin pills or vitamin E (3 × 800 mg) resulted in a significantly smaller increase of DNA effects in WBC of some probands. When the volunteers were given a supplement of vitamin E (1200 mg daily) for 14 days prior to run, exercise-induced DNA damage was clearly reduced in all probands. In four out of five subjects, vitamin supplementation completely prevented the induction of DNA damage after exhaustive exercise. Intake of vitamin E for 14 days led to a clear increase in vitamin E serum concentrations. Malondialdehyde (MDA), a marker of lipid peroxidation, was measured in the serum of probands in tests with and without vitamin supplementation for 14 days. MDA concentrations were significantly decreased following vitamin E supplementation but not significantly changed 15 min and 24 h after a run. Our results demonstrate that vitamin E prevents exercise-induced DNA damage and indicate that DNa breakage occurs in WBC after exhaustive exercise as a consequence of oxidative stress.     

Probucol as a potent inhibitor of oxygen radical-induced lipid peroxidation and DNA damage: in vitro studies

Probucol, a clinically used cholesterol lowering and antioxidant drug, was investigated for possible protection against lipid peroxidation and DNA damage induced by iron nitrilotriacetate (Fe-NTA) plus hydrogen peroxide (H2O2). Fe-NTA is a potent nephrotoxic agent and induces acute and subacute renal proximal tubular necrosis by catalyzing the decomposition of H2O2-derived production of hydroxyl radicals, which are known to cause lipid peroxidation and DNA damage. Fe-NTA is associated with a high incidence of renal adenocarcinoma in rodents. Lipid peroxidation and DNA damage are the principal manifestation of Fe-NTA induced toxicity, which could be mitigated by probucol. Incubation of renal microsomal membrane and/or calf thymus DNA with H2O2 (40 mM) in the presence of Fe-NTA (0.1 mM) induces renal microsomal lipid peroxidation and DNA damage to about 2.4-fold and 5.9-fold, respectively, as compared to control (P < 0.05). Induction of renal microsomal lipid peroxidation and DNA damage was inhibited by probucol in a concentration-dependent manner. In lipid peroxidation protection studies, probucol treatment showed a concentration-dependent inhibition (10-34% inhibition; P < 0.05) of Fe-NTA plus H2O2-induced lipid peroxidation as measured by thiobarbituric acid reacting species' (TBARS) formation in renal microsomes. Similarly, in DNA damage protection studies, probucol treatment also showed a concentration-dependent strong inhibition (36-71% inhibition; P < 0.05) of DNA damage. From these studies, it was concluded that probucol inhibits peroxidation of microsomal membrane lipids and DNA damage induced by Fe-NTA plus H2O2. However, because the lipid peroxidation and DNA damage studied here are regarded as early markers of carcinogenesis, we suggest that probucol may be developed as a cancer chemopreventive agent against renal carcinogenesis and other adverse effects of Fe-NTA exposure in experimental animals, in addition to being a cholesterol-lowering drug, useful for the control of hypercholestrolemia.     

Melatonin reduces Fenton reaction-induced lipid peroxidation in porcine thyroid tissue

Free radicals and reactive oxygen species (ROS) participate in physiological and pathological processes in the thyroid gland. Bivalent iron cation (ferrous, Fe(2+)), which initiates the Fenton reaction (Fe(2+) + H2O2 --> Fe(3+) + *OH + OH(-)) is frequently used to experimentally induce oxidative damage, including that caused by lipid peroxidation. Lipid peroxidation is involved in DNA damage, thus indirectly participating in the early steps of carcinogenesis. In turn, melatonin is a well-known antioxidant and free radical scavenger. The aim of the study was to estimate the effect of melatonin on basal and iron-induced lipid peroxidation in homogenates of the porcine thyroid gland. In order to determine the effect of melatonin on the auto-oxidation of lipids, thyroid homogenates were incubated in the presence of that indoleamine in concentrations of 0.0, 0.00001, 0.0001, 0.001, 0.01, 0.1, 0.25, 0.5, 1.0, 2.5, or 5.0 mM. To study melatonin effects on iron-induced lipid peroxidation, the homogenates were incubated in the presence of FeSO(4) (40 microM) plus H2O2 (0.5 mM), and, additionally, in the presence of melatonin in the same concentrations as above. The degree of lipid peroxidation was expressed as the concentration of malondialdehyde + 4-hydroxyalkenals (MDA + 4-HDA) per mg protein. Melatonin, in a concentration-dependent manner, decreased lipid peroxidation induced by Fenton reaction, without affecting the basal MDA + 4-HDA levels. In conclusion, melatonin protects against iron + H2O2-induced peroxidation of lipids in the porcine thyroid. Thus, the indoleamine would be expected to prevent pathological processes related to oxidative damage in the thyroid, cancer initiation included.     

Protective effects of lazaroid U74389F (16-desmethyl tirilazad) on endrin-induced lipid peroxidation and DNA damage in brain and liver and regional distribution of catalase activity in rat brain

Endrin, a poly-halogenated cyclic hydrocarbon, induces hepatic lipid peroxidation, modulates calcium homeostasis, decreases membrane fluidity, and increases nuclear DNA damage. Little information is available on the neurotoxicity of endrin. The effects of endrin on lipid peroxidation, DNA damage, and regional distribution of catalase activity were assessed in rat brain and liver 24 h following an acute oral dose of 4.5 mg endrin/kg. Lipid peroxidation associated with whole brain mitochondria increased 2.4-fold, whereas microsomal lipid peroxidation increased 2.8-fold following endrin administration. Lipid peroxidation also increased 2.0-fold both in hepatic mitochondria and microsomes. Catalase activity decreased 24% in the hypothalamus, 23% in the cortex, 38% in the cerebellum, and 11% in the brain stem in response to endrin. A 4.3-fold increase in brain nuclear DNA-single strand breaks (SSB) was observed in endrin-treated rats. Pretreatment of rats intraperitoneally with the lazaroid U74389F (16-desmethyl tirilazad) (10 mg/kg in two doses) attenuated the biochemical consequences of endrin-induced oxidative stress. The administration of U74389F in citrate buffer (pH 3.8) provided better protection than administering the lazaroid in corn oil, decreasing endrin-induced lipid peroxidation by 50–80% and DNA-SSB by approximately 72% in liver and 85% in brain, while ameliorating the suppressed catalase activity. The data suggest an involvement of an oxidative stress in the neurotoxicity and hepatotoxicity induced by endrin, which can be attenuated by the lazaroid U74389F.     

Lipid peroxidation-mediated cytotoxicity and DNA damage in U937 cells

Membrane lipid peroxidation processes yield products that may react with DNA and proteins to cause oxidative modifications. In the present study, we evaluated lipid peroxidation-mediated cytotoxicity and oxidative DNA damage in U937 cells. Upon exposure of U937 cells to tert-butylhydroperoxide (t-BOOH) and 2,2'-azobis (2-amidinopropane) hydrochloride (AAPH), which induce lipid peroxidation in membranes, the cells exhibited a reduction in viability and an increase in the endogenous production of reactive oxygen species (ROS), as measured by the oxidation of 2',7'-dichlorodihydrofluorescein. In addition, a significant decrease in the intracellular GSH level and the activities of major antioxidant enzymes were observed. We also observed lipid peroxidation-mediated oxidative DNA damage, reflected by an increase in 8-OH-dG level and loss of the ability of DNA to renature. When the cells were pretreated with the antioxidant N-acetylcysteine (NAC) or the spin trap alpha-phenyl-N-t-butylnitrone (PBN), lipid peroxidation-mediated cytotoxicity in U937 cells was protected. This effect seems to be due to the ability of NAC and PBN to reduce ROS generation induced by lipid peroxidation. These results suggest that lipid peroxidation resulted in a pro-oxidant condition of U937 cells by the depletion of GSH and inactivation of antioxidant enzymes, which consequently leads to a decrease in survival and oxidative damage to DNA. The results indicate that the peroxidation of lipid is probably one of the important intermediary events in oxidative stress-induced cellular damage.     

A new and suitable reconstructed system for NADPH-dependent microsomal lipid peroxidation

In order to evaluate the O-2 participation in NADPH-dependent microsomal lipid peroxidation, we used reconstructed system which contained detergent-solubilized NADPH-dependent cytochrome P-450 reductase, cytochrome P-450, phospholipid liposomes, NADPH and Fe3+-ADP. Lipid peroxidation, monitored by the formation of thiobarbituric acid-reactive substance, was increased with increasing concentration of detergent-solubilized NADPH cytochrome P-450 reductase, cytochrome P-450 or Fe3+-ADP. Cytochrome P-450-dependent lipid peroxidation was parallel to O-2 generation monitored by chemiluminescence probe with 2-methyl-6-(p-methoxyphenol)-3,7-dihydroimidazo[1,2-a]pyrazin++ +-3-one. Lipid peroxidation was significantly inhibited by superoxide dismutase, but not by catalase or sodium benzoate. The reconstructed system herein described is considered to be very close to NADPH-dependent microsomal lipid peroxidation system.     

Importance of UVA photoprotection as shown by genotoxic related endpoints: DNA damage and p53 status

In order to demonstrate the importance of photoprotection in the UVA range (320-400 nm), an in vitro approach where sun formulations are spread on a quartz slide, and placed over human keratinocytes in culture is proposed as a convenient test for photoprotection assessment at the DNA level. Using the comet assay, DNA strand breaks, oxidative DNA damage or drug-induced DNA breaks were assessed. Accumulation of p53 protein was also studied as a marker for UV-induced genotoxic stress. Such a method was used to compare two formulations with different photostability. Spectroradiometry showed that a photounstable formulation lost its effectiveness in UVA screening when pre-irradiated by simulated sunlight (UVB+UVA). As a consequence, it was also shown that this formulation was not as protective as the photostable one at the genomic level. These data demonstrate that the loss of absorbing efficiency within UVA wavelengths due to photounstability may have detrimental consequences leading to impairments implicated in genotoxic events.     

Lipid peroxidation associated cardiolipin loss and membrane depolarization in rat brain mitochondria

Oxidative stress induced by Fe2+ (50 microM) and ascorbate (2 mM) in isolated rat brain mitochondria incubated in vitro leads to an enhanced lipid peroxidation, cardiolipin loss and an increased formation of protein carbonyls. These changes are associated with a loss of mitochondrial membrane potential (depolarization) and an impaired activity of electron transport chain (ETC) as measured by MTT reduction assay. Butylated hydroxytoluene (0.2 mM), an inhibitor of lipid peroxidation, can prevent significantly the loss of cardiolipin, the increased protein carbonyl formation and the decrease in mitochondrial membrane potential induced by Fe2+ and ascorbate, implying that the changes are secondary to membrane lipid peroxidation. However, iron-ascorbate induced impairment of mitochondrial ETC activity is apparently independent of lipid peroxidation process. The structural and functional derangement of mitochondria induced by oxidative stress as reported here may have implications in neuronal damage associated with brain aging and neurodegenerative disorders.