Lycopene inhibits DNA damage and liver necrosis in rats treated with ferric nitrilotriacetate.

Experimental and epidemiological evidence suggests that lycopene, a carotenoid present in tomatoes, tomato products, and several fruits and vegetables, may play a role in preventing certain cancers in humans. We have investigated the effect of lycopene pretreatment on lipid peroxidation, oxidative damage to DNA, and histopathological changes in liver of animals subjected to intraperitoneal (ip) ferric nitrilotriacetate (Fe-NTA) administration. Compared with control rats, liver of Fe-NTA-treated animals showed a significant increase in the 8-oxo-7,8-dihydro-2'-deoxyguanosine level and a 75% increase in malondialdehyde accumulation concomitant with histopathological changes. Five days of lycopene pretreatment (10 mg/kg body weight, ip) almost completely prevented liver biomolecule oxidative damage and protected the tissue against the observed histological alterations.     

DNA damage in digestive gland and mantle tissue of the mussel Perna perna

Data concerning the susceptibility of DNA to damage by reactive oxygen and nitrogen species and other endogenous compounds produced by physiological stress in marine organisms is lacking, especially in bivalve mollusks. In this article, we analyzed the background levels of lipid peroxidation (malondialdehyde, MDA), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and 1,N2-etheno-2'-deoxyguanosine (1,N2-epsilon dGuo) in digestive gland and mantle tissue of mussels Perna perna collected at a cultivation zone in Florianópolis (Santa Catarina, Brazil). The present data point to the possibility of the use of both 8-oxodGuo and 1,N2-epsilon dGuo as complementary indicators of oxidative stress processes in mussels. A sensitive method coupling high performance liquid chromatography to mass spectrometry was applied for the detection of 1,N2-epsilon dGuo in mussel tissues.     

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.     

DNA strand breaks and base modifications induced by cholesterol hydroperoxides

Cholesterol (Ch) can be oxidized by reactive oxygen species, forming oxidized products such as Ch hydroperoxides (ChOOH). These hydroperoxides can disseminate the peroxidative stress to other cell compartments. In this work, the ability of ChOOH to induce strand breaks and/or base modifications in a plasmid DNA model was evaluated. In addition, HPLC/MS/MS analyses were performed to investigate the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) after the incubation of 2'-deoxyguanosine (dGuo) with ChOOH and Cu(2+). In the presence of copper ions, ChOOH induced DNA strand breaks in time and concentration-dependent manners. Purine and pyrimidine base modifications were also observed, as assessed respectively by the treatment with Fpg and Endo III repair enzymes. The detection of 8-oxodGuo by HPLC/MS/MS is in agreement with the dGuo oxidation in plasmid DNA. ChOOH-derived DNA damage adds further support to the role of lipid peroxidation in inducing DNA modifications and mutation.     

DNA strand breaks and base modifications induced by cholesterol hydroperoxides

Abstract

Cholesterol (Ch) can be oxidized by reactive oxygen species, forming oxidized products such as Ch hydroperoxides (ChOOH). These hydroperoxides can disseminate the peroxidative stress to other cell compartments. In this work, the ability of ChOOH to induce strand breaks and/or base modifications in a plasmid DNA model was evaluated. In addition, HPLC/MS/MS analyses were performed to investigate the formation of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) after the incubation of 2′-deoxyguanosine (dGuo) with ChOOH and Cu²⁺. In the presence of copper ions, ChOOH induced DNA strand breaks in time and concentration-dependent manners. Purine and pyrimidine base modifications were also observed, as assessed respectively by the treatment with Fpg and Endo III repair enzymes. The detection of 8-oxodGuo by HPLC/MS/MS is in agreement with the dGuo oxidation in plasmid DNA. ChOOH-derived DNA damage adds further support to the role of lipid peroxidation in inducing DNA modifications and mutation.     

DNA damage and 2'-deoxyguanosine oxidation induced by S(IV) autoxidation catalyzed by copper(II) tetraglycine complexes: synergistic effect of a second metal ion

S(IV) (SO(2),HSO(3)(-)andSO(3)(2-)) autoxidation catalyzed by Cu(II)/tetraglycine complexes in the presence of DNA or 2'-deoxyguanosine (dGuo) resulted in DNA strand breaks and formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), respectively. Ni(II), Co(II) or Mn(II) (1.0x10(-4)M) complexes had much smaller effects. Cu(II)/tetraglycine (1.0x10(-4)M) in the presence of Ni(II) or Mn(II) (10(-7)-10(-6)M) and S(IV) showed remarkable synergistic effect with these metal ions producing a higher yield of 8-oxodGuo. Oxidation of dGuo and DNA damage were attributed to oxysulfur radicals formed as intermediates in S(IV) autoxidation catalyzed by transition metal ions. SO*(3)(-) and HO* radicals were detected by EPR-spin trapping experiments with DMPO (5,5-dimethyl-1-pyrroline-N-oxide).     

Experimental approaches to free radicals and ageing

Abstract

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.     

Chemical and biological consequences of oxidatively damaged guanine in DNA

Of the four native nucleosides, 2'-deoxyguanosine (dGuo) is most easily oxidized. Two lesions derived from dGuo are 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (Fapy)?dGuo. Furthermore, while steady-state levels of 8-oxodGuo can be detected in genomic DNA, it is also known that 8-oxodGuo is more easily oxidized than dGuo. Thus, 8-oxodGuo is susceptible to further oxidation to form several hyperoxidized dGuo products. This review addresses the structural impact, the mutagenic and genotoxic potential, and biological implications of oxidatively damaged DNA, in particular 8-oxodGuo, Fapy?dGuo, and the hyperoxidized dGuo products.     

Glutathione metabolizing enzymes and oxidative stress in ferric nitrilotriacetate mediated hepatic injury

Summary

Glutathione (GSH) plays several important roles in the protection of cells against oxidative damage, particularly following exposure to xenobiotics. Ferric nitrilotriacetate (Fe-NTA) is a potent depletor of GSH and also enhances tissue lipid peroxidation. In this study, we show the effect of Fe-NTA treatment on hepatic GSH and some of the glutathione metabolizing enzymes, oxidant generation and liver damage. The level of hepatic GSH and the activities of glutathione reductase, glutathione S-transferase, glutathione peroxidase, and glucose 6-phosphate dehydrogenase all decrease following Fe-NTA administration. In these parameters the maximum decrease occurred at 12 h following Fe-NTA treatment. In contrast, γ-glutamyl transpeptidase was increased at this time. Not surprisingly, the increase in the activity of γ-glutamyl transpeptidase and decreases in GSH, glutathione peroxidase, glutathione reductase, glucose 6-phosphate dehydrogenase and glutathione S-transferase were found to be dependent on the dose of Fe-NTA administered. Fe-NTA administration also enhances the production of H₂O₂ and increases hepatic lipid peroxidation. Parallel to these changes, Fe-NTA enhances liver damage as evidenced by increases in serum transaminases. Once again, the liver damage is dependent on the dose of Fe-NTA and is maximal at 12 h. Pretreatment of animals with antioxidant, butylated hydroxy anisole (BHA), protects against Fe-NTA-mediated hepatotoxicity further supporting the involvement of oxidative stress in Fe-NTA-mediated hepatic damage. In aggregate, our results indicate that Fe-NTA administration eventuates in decreased hepatic GSH, a fall in the activities of glutathione metabolizing enzymes and excessive production of oxidants, all of which are involved in the cascade of events leading to iron-mediated hepatic injury.     

Chemical and biological consequences of oxidatively damaged guanine in DNA

Abstract

Of the four native nucleosides, 2′-deoxyguanosine (dGuo) is most easily oxidized. Two lesions derived from dGuo are 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (Fapy)?dGuo. Furthermore, while steady-state levels of 8-oxodGuo can be detected in genomic DNA, it is also known that 8-oxodGuo is more easily oxidized than dGuo. Thus, 8-oxodGuo is susceptible to further oxidation to form several hyperoxidized dGuo products. This review addresses the structural impact, the mutagenic and genotoxic potential, and biological implications of oxidatively damaged DNA, in particular 8-oxodGuo, Fapy?dGuo, and the hyperoxidized dGuo products.     

Simultaneous determination of O6-methyl-2'-deoxyguanosine, 8-oxo-7,8-dihydro-2'-deoxyguanosine, and 1,N6-etheno-2'-deoxyadenosine in DNA using on-line sample preparation by HPLC column switching coupled to ESI-MS/MS

O(6)-Methyl-2'-deoxyguanosine (O(6)-mdGuo), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), and 1,N(6)-etheno-2'-deoxyadenosine (epsilondAdo) are promutagenic DNA lesions originating from both endogenous and exogenous agents and actions (methylation, hydroxylation, lipid peroxidation products). A highly sensitive quantitative method was developed to measure these DNA adducts simultaneously, using liquid chromatography tandem mass spectrometry with column switching. Deuterated O(6)-[(2)H(3)]mdGuo was synthesized and used as internal standard. The limits of quantification for O(6)-mdGuo, 8-oxodGuo, and epsilondAdo were 24, 98, and 48 fmol on column, respectively. The method showed linearity in the range 0.24-125 pmol/ml, 0.98-125 pmol/ml, and 0.49-62.5 pmol/ml for the three adducts, respectively. The inter-day precision in the linear concentration range was between 1.7 and 9.3% for O(6)-mdGuo, 10.6 and 28.7% for 8-oxodGuo, and 6.2 and 10.4%, for epsilondAdo. In DNA isolated from liver of untreated 12-week-old female F344 rats, O(6)-mdGuo was above the limit of detection (37 adducts per 10(9) normal nucleosides) but could not be quantified. 8-oxodGuo and epsilondAdo showed background levels of 500 and 130 adducts per 10(9) normal nucleosides, respectively. DNA analyzed 1h after treatment of rats with dimethylnitrosamine by oral gavage of 50 microg/kg b.wt. did not affect the levels of 8-oxodGuo and epsilondAdo but resulted in 200 O(6)-mdGuo adducts per 10(9) normal nucleosides. The method developed will be of use to study the biological significance of exogenous DNA adducts as an increment to background DNA damage and the role of modulating factors, such as DNA repair.     

Strobilanthes crispus attenuates renal carcinogen, iron nitrilotriacetate (Fe-NTA)-mediated oxidative damage of lipids and DNA

This study was aimed to evaluate the effect of Strobilanthes crispus extract for possible protection against lipid peroxidation and DNA damage induced by iron nitrilotriacetate (Fe-NTA) and 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. Incubation of postmitochondrial supernatant and/or calf thymus DNA with H₂O₂ (40 mM) in the presence of Fe-NTA (0.1 mM) induces lipid peroxidation and DNA damage to about 2.3-fold and 2.9-fold, respectively, as compared to control (P < 0.05). In lipid peroxidation protection studies, S. crispus treatment showed a dose-dependent inhibition (45–53% inhibition, P < 0.05) of Fe-NTA and H₂O₂ induced lipid peroxidation. Similarly, in DNA damage protection studies, S. crispus treatment also showed a dose-dependent inhibition (18–30% inhibition, P < 0.05) of DNA damage. In addition, the protection was closely related to the content of phenolic compounds as evident by S. crispus extract showing the value of 124.48 mg/g total phenolics expressed as gallic acid equivalent (GAE, mg/g of extract). From these studies, it is concluded that S. crispus inhibits peroxidation of membrane lipids and DNA damage induced by Fe-NTA and H₂O₂ and possesses the potential to be used to treat or prevent degenerative diseases where oxidative stress is implicated.     

Antioxidant ebselen reduces oxidative damage in focal cerebral ischemia

The antioxidant and neuroprotective potential of the glutathione peroxidase mimic ebselen has been investigated in experimental stroke. Intravenous ebselen (1 mg/kg/h) or vehicle infusion was started 45 min before permanent middle cerebral artery occlusion in the rat, and continued until the end of the experiment. The topography and extent of oxidative damage to the brain was assessed immunohistochemically using an antibody for DNA damage that identified hydroxylated products of 2'-deoxyguanosine (8-OHdG/8-oxodGuo) and an antibody for lipid peroxidation that identified the 4-hydroxynonenal histidine adduct (4-HNE). Ischemic damage was mapped and evaluated with standard histopathology. In the vehicle-treated rats immunopositive staining for both 8-oxodGuo and 4-HNE extended beyond the boundary of ischemic damage. In ebselen-treated rats, the extent of tissue immunopositive for 8-oxodGuo, and 4-HNE was less than that demonstrating ischemic damage confirming the antioxidant mechanism of action in vivo. In addition, ebselen treatment induced a 28% reduction in cortical ischemic damage (p <.02).     

Glyceryl trinitrate,a nitric oxide donor,abrogates ferric nitrilotriacetate-induced oxidative stress and renal damage

Ferric nitrilotriacetate (Fe-NTA), a common water pollutant and a known renal carcinogen, acts through the generation of oxidative stress and hyperproliferative response. In the present study, we show that the nitric oxide (NO) generated by the administration of glyceryl trinitrate (GTN) affords protection against Fe-NTA-induced oxidative stress and proliferative response. Administration of Fe-NTA resulted in a significant (P<0.001) depletion of renal glutathione (GSH) content with concomitant increase in lipid peroxidation and elevated tissue damage marker release in serum. Parallel to these changes, Fe-NTA also caused down-regulation of GSH metabolizing enzymes including glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione-S-transferase and several fold induction in ornithine decarboxylase (ODC) activity and rate of DNA synthesis. Subsequent exogenous administration of GTN at doses of 3 and 6mg/kg body weight resulted in significant (P<0.001) recovery of GSH metabolizing enzymes and amelioration of tissue GSH content, in a dose-dependent manner. GTN administration also inhibited malondialdehyde (MDA) formation, induction of ODC activity, enhanced rate of DNA synthesis, and pathological deterioration in a dose-dependent fashion. Further, administration of NO inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME), exacerbated Fe-NTA-induced oxidative tissue injury, hyperproliferative response, and pathological damage. Overall, the study suggests that NO administration subsequent to Fe-NTA affords protection against ROS-mediated damage induced by Fe-NTA.     

Protective role of zinc-metallothionein (Zn-MT) in iron nitrilotriacetate (Fe-NTA)-induced renal oxidative damage

Several studies have shown the role of thiol-rich proteins especially metallothionein (MT) in the therapeutic interventions against oxidative damage. Previously, we have provided strong evidence for the involvement of ROS in iron nitrilotriacetate (Fe-NTA)-induced renal toxicity, which may have relevance to its carcinogenicity. The purpose of this study was to evaluate the role of zinc metallothionein (Zn-MT) on the protection against Fe-NTA-induced renal oxidative damage. The results demonstrate that Zn-MT pretreatment provided protection against Fe-NTA-induced mortality in mice (40% protection). Similarly, Zn-MT pretreatment also provided protection against Fe-NTA-induced lipid peroxidation (26% inhibition, P < 0.001). It is proposed that Zn-MT protects kidney tissue against the noxious effect of Fe-NTA primarily by interference with lipid peroxides. It is concluded that Zn-MT may serve as an excellent physiological antioxidant against Fe-NTA-mediated renal oxidative damage.     

Analysis of monohydroxyeicosatetraenoic acids and F2-isoprostanes as markers of lipid peroxidation in rat brain mitochondria

We have introduced two specific techniques for the quantitative measurement of monohydroxyeicosatetraenoic acids (HETEs) and F2-isoprostanes by gas chromatography-mass spectrometry/negative ion chemical ionization (GC-MS/NICI) to study lipid peroxidation in isolated rat brain mitochondria by iron/ascorbate. The analysis of HETEs involved hydrogenation, solid phase extraction on a C18-cartridge, formation of pentafluorobenzyl bromide and trimethylsilyl ether derivatives. In the case of F2-isoprostanes, the analytical procedure was similar to that of HETEs except that the hydrogenation step was omitted. We found that HETE content (sum of 5-, 8-12-, and 15-isomers) in freshly prepared rat brain mitochondria was 220 +/- 40pmol/mg protein. The corresponding content for the F2-isoprostane, 8-iso-PGF2alpha, was 0.21 +/-+/- 0.10 pmol/mg protein. HETEs and 8-iso-PGF2alpha were predominantly present in the esterified form. The content of both HETEs and 8-iso-PGF2alpha were increased in presence of iron/ascorbate as oxidation system. After 30 min incubation with Fe2+ ascorbate, the content of HETE isomers was increased about 6-fold compared with baseline levels whereas that for 8-iso-PGF2alpha was elevated 100-fold. Formation of HETEs and F2-isoprostanes corresponded to the consumption of arachidonic acid (AA) and alpha-tocopherol, respectively. There were almost no changes in the content of free (non-esterified) HETEs and 8-iso-PGF2alpha during the course of iron/ascorbate induced oxidation of the brain mitochondria. Our data provide the first direct evidence for the presence of HETEs and F2-isoprostanes in freshly isolated rat brain mitochondria and that esterified HETEs and 8-iso-PGF2alpha are predominantly generated during iron/ascorbate induced lipid peroxidation. Sensitive quantification of these products of non-enzymatic lipid peroxidation as indicators of oxidant injury opens new areas of investigation regarding the role of free radicals in the pathogenesis of human diseases. In addition, HETEs and F2-isoprostanes may be important mediators for mitochondrial functions.     

Carotenoid-containing unilamellar liposomes loaded with glutathione: a model to study hydrophobic-hydrophilic antioxidant interaction

Unilamellar liposomes are used as a simple two-compartment model to study the interaction of antioxidants. The vesicle membrane can be loaded with lipophilic compounds such as carotenoids or tocopherols, and the aqueous core space with hydrophilic substances like glutathione (GSH) or ascorbate, mimicking the interphase between an aqueous compartment of a cell and its surrounding membrane.

Unilamellar liposomes were used to investigate the interaction of GSH with the carotenoids lutein, β-carotene and lycopene in preventing lipid peroxidation. Lipid peroxidation was initiated with 2,2′-azo-bis-[2,4-dimethylvaleronitrile] (AMVN). Malondialdehyde (MDA) formation was measured as an indicator of oxidation; additionally, the loss of GSH was followed. In liposomes without added antioxidant, MDA levels of 119 ± 6 nmol/mg phospholipid were detected after incubation with AMVN for 2 h at 37°C. Considerably lower levels of 57 ± 8 nmol MDA/mg phospholipid were found when the liposomal vesicles had been loaded with GSH. Upon incorporation of β-carotene, lycopene or lutein, the resistance of unilamellar liposomes towards lipid peroxidation was further modified. An optimal further protection was observed with 0.02 nmol β-carotene/mg phospholipid or 0.06 nmol lycopene/mg phospholipid. At higher levels both these carotenoids exhibited prooxidant effects. Lutein inhibited lipid peroxidation in a dose-dependent manner between 0.02 and 2.6 nmol/mg phospholipid. With increasing levels of lycopene and lutein the consumption of encapsulated GSH decreased moderately, and high levels of β-carotene led to a more pronounced loss of GSH.

The data demonstrate that interactions between GSH and carotenoids may improve resistance of biological membranes towards lipid peroxidation. Different carotenoids exhibit specific properties, and the level for optimal protection varies between the carotenoids.     

Vitamin E inhibits hepatic oxidative stress, toxicity and hyperproliferation in rats treated with the renal carcinogen ferric nitrilotriacetate

Ferric nitrilotriacetate (Fe-NTA) is a potent renal and hepatic tumor promoter, which acts through a mechanism involving oxidative stress. Fe-NTA when injected intraperitoneally into rats induces hepatic ornithine decarboxylase activity as well as hepatic DNA synthesis. Vitamin E is a well-known, lipid-soluble and chain-breaking antioxidant which protects cell membranes from peroxidative damage. In this study, we investigated the protective effect of vitamin E, a major fat-soluble antioxidant, against Fe-NTA-mediated hepatic oxidative stress, toxicity and hyperproliferation in Wistar rats. Animals were treated with two different doses of vitamin E for 1 week prior to Fe-NTA treatment. Vitamin E at a higher dose of 2.0 mg/animal/day showed significant reduction in Fe-NTA-induced hepatic ornithine decarboxylase activity, DNA synthesis, microsomal lipid peroxidation and hydrogen peroxide generation. Fe-NTA treatment alone caused depletion of glutathione, glutathione metabolizing and antioxidant enzymes in rat liver, whereas pretreatment of animals with vitamin E reversed these changes in a dose-dependent manner. Taken together, our results suggest that vitamin E may afford substantial protection against the damage caused by Fe-NTA exposure and can serve as a potent preventive agent to suppress oxidant-induced tissue injury.     

Carotenoid-containing unilamellar liposomes loaded with glutathione: a model to study hydrophobic-hydrophilic antioxidant interaction

Unilamellar liposomes are used as a simple two-compartment model to study the interaction of antioxidants. The vesicle membrane can be loaded with lipophilic compounds such as carotenoids or tocopherols, and the aqueous core space with hydrophilic substances like glutathione (GSH) or ascorbate, mimicking the interphase between an aqueous compartment of a cell and its surrounding membrane.

Unilamellar liposomes were used to investigate the interaction of GSH with the carotenoids lutein, β-carotene and lycopene in preventing lipid peroxidation. Lipid peroxidation was initiated with 2,2'-azo-bis-[2,4-dimethylvaleronitrile] (AMVN). Malondialdehyde (MDA) formation was measured as an indicator of oxidation; additionally, the loss of GSH was followed. In liposomes without added antioxidant, MDA levels of 119 ± 6 nmol/mg phospholipid were detected after incubation with AMVN for 2 h at 37°C. Considerably lower levels of 57 ± 8 nmol MDA/mg phospholipid were found when the liposomal vesicles had been loaded with GSH. Upon incorporation of β-carotene, lycopene or lutein, the resistance of unilamellar liposomes towards lipid peroxidation was further modified. An optimal further protection was observed with 0.02 nmol β-carotene/mg phospholipid or 0.06 nmol lycopene/mg phospholipid. At higher levels both these carotenoids exhibited prooxidant effects. Lutein inhibited lipid peroxidation in a dose-dependent manner between 0.02 and 2.6 nmol/mg phospholipid. With increasing levels of lycopene and lutein the consumption of encapsulated GSH decreased moderately, and high levels of β-carotene led to a more pronounced loss of GSH.

The data demonstrate that interactions between GSH and carotenoids may improve resistance of biological membranes towards lipid peroxidation. Different carotenoids exhibit specific properties, and the level for optimal protection varies between the carotenoids.     

Counterpoint,patronage and professionalism

Abstract

Several studies have shown the role of thiol-rich proteins especially metallothionein (MT) in the therapeutic interventions against oxidative damage. Previously, we have provided strong evidence for the involvement of ROS in iron nitrilotriacetate (Fe-NTA)-induced renal toxicity, which may have relevance to its carcinogenicity. The purpose of this study was to evaluate the role of zinc metallothionein (Zn-MT) on the protection against Fe-NTA-induced renal oxidative damage. The results demonstrate that Zn-MT pretreatment provided protection against Fe-NTA-induced mortality in mice (40% protection). Similarly, Zn-MT pretreatment also provided protection against Fe-NTA-induced lipid peroxidation (26% inhibition, P < 0.001). It is proposed that Zn-MT protects kidney tissue against the noxious effect of Fe-NTA primarily by interference with lipid peroxides. It is concluded that Zn-MT may serve as an excellent physiological antioxidant against Fe-NTA-mediated renal oxidative damage.