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Alcoholic patients and experimental animals exposed to ethanol display biochemical signs of oxidative damage, suggesting a possible role of free radicals in causing some of the toxic effects of alcohol. The use of electron spin resonance (ESR) spectroscopy associated with spin trapping technique has demonstrated that hydroxyethyl radicals are generated during ethanol metabolism by the microsomal monoxygenase system, involving the alcohol-inducible cytochrome P450 2E1 (CYP2E1). Recent observations in rats fed intragastrically with a high fat diet containing ethanol indicate that the formation of hydroxyethyl radicals is associated with stimulation of lipid peroxidation and development of liver damage. Moreover, by alkylating liver proteins, and particularly CYP2E1, hydroxyethyl free radicals are also capable of inducing the production of specific antibodies which can be observed in ethanol-fed animals as well as in patients abusing alcohol. Recent studies have demonstrated that hydroxyethyl radical-derived antigens are exposed on the plasma membranes of hepatocytes exposed to ethanol where are able to target antibody-dependent cell-mediated immunotoxic reactions towards liver cells. Thus, beside to contribute to alcohol-mediated oxidative damage, hydroxyethyl free radicals can contribute by immunological mechanisms to cause hepatocellular lesions associated with alcohol abuse.

Objectives: Laboratory studies suggest that myocardial reperfusion injury is exacerbated by free radicals when pure oxygen is used during cardiopulmonary bypass. In phase I of this study we demonstrated that normoxic perfusion during cardiopulmonary bypass does not increase the risk of microembolic brain injury so long as a membrane oxygenator with an arterial filter is used. In phase II of this study we studied the hypothesis that normoxic perfusion increases the risk of hypoxic brain injury after deep hypothermia with circulatory arrest., Methods: With membrane oxygenators with arterial filters, 10 piglets (8-10 kg) underwent 120 minutes of deep hypothermia and circulatory arrest at 15 degrees C, were rewarmed to 37 degrees C, and were weaned from bypass. In 5 piglets normoxia (PaO2 64-181 mm Hg) was used during cardiopulmonary bypass and in 5 hyperoxia (PaO2 400-900 mm Hg) was used. After 6 hours of reperfusion the brain was fixed for histologic evaluation. Near-infrared spectroscopy was used to monitor cerebral oxyhemoglobin and oxidized cytochrome a,a3 concentrations., Results: Histologic examination revealed a significant increase in brain damage in the normoxia group (score 12.4 versus 8.6, P =.01), especially in the neocortex and hippocampal regions. Cytochrome a,a 3 and oxyhemoglobin concentrations tended to be lower during deep hypothermia and circulatory arrest in the normoxia group (P =.16)., Conclusions: In the setting of prolonged deep hypothermia and circulatory arrest with membrane oxygenators, normoxic cardiopulmonary bypass significantly increases histologically graded brain damage with respect to hyperoxic cardiopulmonary bypass. Near-infrared spectroscopy suggests that the mechanism is hypoxic injury, which presumably overwhelms any injury caused by increased oxygen free radicals.

Angiotensin-converting enzyme inhibitors have been demonstrated to protect spontaneously hypertensive rats from cerebral ischemia. The present study investigated the protective effect of enalapril and moexipril in models of permanent focal cerebral ischemia in normotensive mice and rats. To elucidate the mechanism of neuroprotection the influence of these angiotensin-converting enzyme inhibitors on glutamate-, staurosporine- or Fe2+/3+-induced generation of reactive oxygen species and neuronal cell death in primary cultures from chick embryo telencephalons was studied. Treatment with moexipril or enalapril dose-dependently reduced the percentage of damaged neurons, as well as mitochondrial reactive oxygen species generation induced by glutamate, staurosporine or Fe2+/3+. Furthermore, moexipril and enalapril attenuated staurosporine-induced neuronal apoptosis as determined by nuclear staining with Hoechst 33258. In mice, 1 h pretreatment with enalapril (0.03 mg/kg) or moexipril (0.3 mg/kg) significantly reduced brain damage after focal ischemia as compared to control animals. Additionally, moexipril (0.01 mg/kg) was able to reduce the infarct volume in the rat model after focal cerebral ischemia. The results of the present study indicate that the angiotensin-converting enzyme inhibitors enalapril and moexipril promote neuronal survival due to radical scavenging properties.

Recent work with a guinea pig model of otitis media has demonstrated evidence of oxygen free radical damage to the middle ear mucosa. However, the relevance of an animal model to human disease is uncertain. Accordingly, the following pilot study was conducted to examine human middle ear fluid for lipid hydroperoxides as evidence of free radical damage. Thirty-five specimens of middle ear fluid from children with chronic otitis media were collected and described as mucoid (n = 19), purulent (n = 10), or serous (n = 6); specimens were weighed and analyzed for lipid hydroperoxide content. The results demonstrated the presence of lipid hydroperoxide in all 3 types of middle ear fluid. Additionally, there was a statistically significant elevation of total lipid hydroperoxide content in mucoid effusions compared with serous effusions, as well as a significant elevation of lipid hydroperoxide divided by weight of purulent effusions compared with serous effusions. This is the first study to document free radical damage in human otitis media.

Kwashiorkor remains a significant cause of childhood morbidity and mortality in tropical regions. However, there are relatively few reproducible animal models to elucidate the aetiopathogenesis, biochemical derangements, and morphological changes of this condition. In the present study, 64 weanling male Wistar albino rats were divided into high protein chow, high protein maize starch, and low protein maize starch groups. After 4 weeks on these respective diets, animals on the low protein diet were commenced on the high protein maize starch diet, while those in the other groups were continued on their original diets for a further period of 5 weeks each The low protein group of animals initially showed growth failure, hair loss, oedoma, fatty liver, metabolic acidosis, hypoalbuminaemia, and anaemia, which are characteristic features of human kwashiorkor. These changes were all reversed by dietary rehabilitation with the high protein diet. The reduced plasma retinol, increased plasma alanine and aspartate aminotransferases and fatty liver observed in the low protein group of animals probably suggest an important role for free radicals in the aetiopathogenesis of kwashiorkor.

The two most striking features of Alzheimer's disease are: a) the multitude of abnormalities affecting essentially every system, and b) the strict age dependence. Recent work suggests that both features are linked to increased oxidative stress that damages lipids, proteins and nucleic acids and results in redox-active metal accumulations, mitochondrial damage and formation of advanced glycation endproducts. Interestingly, beta-protein precursor, amyloid-beta, presenilins and apolipoprotein E have all been linked to reactive oxygen species production or with apoptosis, a process intimately associated with oxidative stress. In therapeutics, the commonality between a number of efficacious agents appears to be oxidative stress reduction. Therefore, we contend that oxidative stress is the element that links the multitude of changes in Alzheimer's disease and that a reduction of oxidative stress will have a dramatic effect on reducing the incidence or progression of Alzheimer's disease.

Mounting clinical and experimental evidence indicates that free radicals play important roles in many physiological and pathological conditions. The wider application of free radical measurement has increased awareness of functional implications of radical-induced impairment of the oxidative/antioxidative balance. In the following review, the role of oxygen free radicals in some human and experimental pathological conditions is described, with particular emphasis on the mechanisms by which they produce oxidative damage to lipids, proteins, and nucleic bases. The role of free radicals and the activation of the antioxidant systems in arteriosclerosis and ageing, diabetes, ischemia/reperfusion injury, ethanol intoxication, and liver steatosis is discussed. Therapeutic approaches to the use of antioxidants have been described and prospects for clinical use have been considered.

Congenital malformations remain the major cause of morbidity and mortality among the offspring of women with diabetes. Animal and human studies indicate that these anomalies occur very early in pregnancy and result from derangements of the maternal metabolic fuels which support embryogenesis. The mechanism for induction of dysmorphogenesis in experimental diabetic pregnancy has been shown to include deficiency states of membrane lipids (myoinositol, arachidonic acid, etc.), alteration in the prostaglandin cascade, and the generation of excess free oxygen radicals. These biochemical alterations result in characteristic morphological and molecular changes which are considered to be the basis of diabetic embryopathy. This article not only discusses the pathomechanism, but also reviews both clinical and experimental strategies to prevent diabetes-associated birth defects.

Recent studies have shown that synthase for nitric oxide or scavenger enzymes is distributed throughout the endometrium. We have reported that endothelial nitric oxide synthase, originally identified in vascular endothelial cells, is distributed in glandular epithelial cells in the endometrium, peaking in the midsecretory phase. In addition, it is known that superoxide dismutase is distributed throughout the endometrium, varying with the menstrual cycle. Yet it is not clear how these enzymes are committed in the reproductive processes. Endometriosis is often complicated by infertility and miscarriage. Of particular interest is that these enzymes are overexpressed in the disease throughout the menstrual cycle. These findings strongly suggest that excessive amounts of free radicals are produced in endometriosis. Copyrightz1999S.KargerAG,Basel

The skin is supplied with an antioxidant system that includes enzymatic and nonenzymatic components. This complex system is the skin's first-line defense against free-radical attacks. Antioxidants are essential in protecting the epidermis from damage by free radicals generated both by environmental and endogenous factors. This role of free radicals in etiology of disease and how antioxidants are used to offset or prevent oxidative damage are discussed.

Experimental studies and few human reports demonstrate that hyperoxia increases the level of reactive oxygen-derived free radicals and that these substances can produce oxidative cellular injury. However, available data suggest that the human lung is more resistant to hyperoxic oxidative damage than previously expected and demonstrate that absorption atelectasis is the most frequently pulmonary effect of inhalation of a high inspired oxygen fraction. Practically, the therapeutic use of high inspired oxygen fractions is limited to patients with acute lung injury and severe hypoxemia. Recent studies demonstrated that ventilator-induced lung injury is a more important cause of pulmonary damage in these patients than hyperoxic toxicity. New protective ventilatory strategies are associated with increased survival.

Aim: To test the hypothesis that complications of neonatal intensive care are related to increased oxygen derived free radical activity, using breath pentane as a marker of lipid peroxidation., Methods: Exhaled breath was collected daily from 57 ventilated preterm infants and pentane concentration measured by gas chromatography., Results: High peak pentane exhalation was significantly associated with low gestational age, mortality, intraventricular haemorrhage and retinopathy of prematurity. Peak pentane was not significantly associated with the development of chronic lung disease., Conclusions: The demonstration that pentane exhalation is related to the course of neonatal disease and its outcome is consistent with the hypothesis that lipid peroxidation is associated with these illnesses, and may contribute to their severity. If this is a causal relation, antioxidant treatments could prove useful in reducing their severity. Measurement of breath pentane might assist in the assessment of antioxidant strategies prior to more extensive clinical trials.

Recent studies have identified several factors which may affect human health and life expectancy in northern Finland. They have shown that antioxidants, infections, genetic or environmental factors may affect the development of and morbidity/mortality from cardiovascular diseases, cancer, diabetes mellitus and other diseases in the northern provinces of this country. Both the occurrence and mortality from coronary heart disease (CHD) is low in the northernmost part of the country, i.e. Mountain Lapland or the Saami area, compared with that in whole country or a neighbouring region to the south in central Lapland. The mortality from all diseases is also low in communities in Mountain Lapland, and high in central Lapland in communities such as Kittilä and Kolari. High scrum antioxidants, alpha-tocopherol (vitamin E), albumin and selenium levels have been measured in men living in the northernmost part of the country, where the death rate from CHD is low. Low serum alpha-tocopherol and albumin levels were typical of men living in rural communities with high CHD mortality, e.g. Kittilä community. Serum antioxidant levels were related to the diet; alpha-tocopherol increased with the consumption of reindeer meat and selenium with fish consumption. Our earlier studies have also identified a low Chlamydia pneumoniae IgA antibody titer in men living in Mountain Lapland compared with men in the neighboring region to the south in central Lapland with high CHD mortality. An elevated Chlamydia pneumoniae IgA antibody titer was associated with low serum alpha-tocopherol level. The people of Saami origin, an ethnic minority living in northernmost Finland, have a high apolipoprotein (apo) E e4 allele frequency and high serum cholesterol. They also have more apo A-IV-2 allele than most of the studied populations, and their HDL cholesterol levels are higher in apo A-IV-2/1 than in apo A-IV-1/1 phenotypes. Our earlier studies indicate that people living in northeastern Finland, west of smelters in Kola Peninsula may be exposed to heavy metals such as cadmium and mercury. Blood cadmium was related to blood pressure and high in men with arterial hypertensive disease. The investigations presented in this article form a good basis for further studies that clarify underlying reasons the health problems in the north.

The acute phase of reflex sympathetic dystrophy (RSD) is characterized by the classical signs and symptoms of inflammation (rubor, calor, dolor, tumor and impaired function). As free radicals are involved in acute inflammation, we studied the effects of free radicals in an animal model, especially as to signs and symptoms found in acute RSD. Awake rats were given continuous intra-arterial infusion (1 ml/h) in the left hindlimb, with saline (n = 6) or the free-radical donor tert-butylhydroperoxide (tert-BuOOH, 25 mM, n = 6). During a 24-h infusion period the skin temperature, volume, skin color, function and pain reactions of the paws were observed. After 24 h the rats were killed and both gastrocnemius muscles were histologically analyzed. Infusion with tert-BuOOH induced in the left paw an increased skin temperature, increased volume, redness of the plantar skin, impaired function and increased pain sensation, while these acute RSD signs and symptoms were absent in the saline infused animals. The alterations in pain sensation (spontaneous, mechanical and thermal pain) were similar to findings in the neuropathic animal model. The gastrocnemius muscles of the saline infused rats and the contralateral gastrocnemius muscle of the tert-BuOOH infused rats showed no histological tissue damage. In the left gastrocnemius muscle free-radical-related damage was visible. Induction of free-radical formation in one hindlimb of awake rats mimics the acute signs and symptoms of acute RSD, with alterations in pain sensation as found in the classical neuropathic animal model of RSD, as well as in acute RSD patients.

In the present study the effect of ascorbate (0.8 mM)/iron (2.5 microM) on lipid and protein oxidation, in Synaptosomes isolated from rat brain cortex, was evaluated. Vitamin E, idebenone and reduced glutathione were used as free radicals scavengers, in order to analyze the mechanism involved in ascorbate/iron-induced oxidative stress. An increased formation of reactive oxygen species (ROS) in the cytosol and in the mitochondria was observed, in ascorbate/iron treated synaptosomes. Idebenone (50 microM) prevented the increased formation of ROS in both synaptosomal compartments, vitamin E (150 microM) protected partially this formation in mitochondria, whereas reduced glutathione (250 microM) (GSH) was ineffective. After ascorbate/iron treatment an increase in lipid peroxidation occurred as compared to control, which was completely inhibited by idebenone. A decrease in protein-SH content was also observed, and it was prevented by Vitamin E, idebenone and GSH. When synaptosomes were treated with ascorbate/iron the levels of GSH decreased, and the levels of oxidized glutathione (GSSG) increased as compared to controls under these conditions. Glutathione peroxidase activity was unchanged, whereas an inhibition of glutathione reductase activity was observed. These data suggest that the increased formation of free radicals in synaptosomes leads to lipid and protein oxidation, the role of the endogenous GSH being essential to protect protein thiol-groups against oxidative damage in order to maintain enzyme activity.

A selenium-containing catalytic antibody (Se-4A4), prepared by converting reactive serine residues of a monoclonal antibody (4A4) raised against a GSH derivative into selenocysteines, acts as a mimic of cytosolic glutathione peroxidase (cGPX). To clarify the mechanism of action of this catalytic antibody, detailed studies on kinetic behaviour and biological activity were carried out. A rate of acceleration (kcat/Km/kuncat) 10(7)-fold that of the uncatalytic reaction is observed. Under similar conditions, the turnover number (kcat) of Se-4A4 is 42% of that of the natural rabbit liver cGPX. The Se-4A4 reaction involves a Ping Pong mechanism, which is the same as that of the natural cGPX. The selenocysteine residue is located in the binding site of the antibody and is shown to be crucial for this activity. Of the thiol compounds tested, only GSH is able to serve as substrate for Se-4A4. It was demonstrated, using the free-radical-damage system (hypoxanthine/xanthine oxidase) of cardiac mitochondria, that Se-4A4 can protect mitochondria from free-radical damage at least 10(4)-fold more effectively than the natural cGPX.

Epidemiological evidence has revealed an inverse relationship such that increased physical activity as measured directly subjective recall, job classification, former athletic status, or indirectly by physical fitness is associated with decreased incidence and (or) mortality rates for various cancers. The relationship appears strongest for colon cancer and female estrogen-dependent cancers of the breast, ovary, and endometrium. While some epidemiological studies have controlled for numerous confounding variables such as smoking, body mass index, and percent body fat, it is still difficult to ascertain whether physical activity exerts an independent effect on cancer above and beyond that associated with an improved lifestyle and numerous other potential confounding variables. Experimental studies performed in animals suggests that chronic exercise, especially when performed prior to tumorigenesis, can retard, delay, or prevent the incidence, progression, or spread of experimental tumors. There is also limited animal evidence suggesting that exercise may help ameliorate cancer cachexia. Exercise or physical activity may contribute to a reduction in site-specific cancers by different physiological mechanisms. Some purported mechanisms include decreased lifetime exposure to estrogen or other hormones, reduced body fat, enhanced gut motility, improved anti-oxidant defenses, and stimulation of anti-tumor immune defenses. Unfortunately, most animal studies have failed to account for plausible biological mechanisms as to how exercise might influence cancer. In addition, the exercise or activity dosage required to provide optimal protection from cancer is unclear. Interpretation of epidemiological studies is hampered by the numerous and sometimes inaccurate assessments of physical activity. Likewise, many animal studies have utilized unrealistic exercise protocols. Clearly, more research is needed to define appropriate activity or exercise dosages definitively and to explore the mechanism(s) by which exercise helps protect against cancer. Nevertheless, moderate exercise appears to be a safe and effective means of aiding in the prevention of cancer and should be adopted by the public in addition to other prudent behavioral practices such as proper diet. More research needs to be performed regarding the effects of exercise or physical activity on those who already have cancer to determine if exercise improves their prognosis.

Oxidative injury caused by free radicals is an important cause of tissue injury now recognized to occur in inflammation, ischemia and by the action of xenobiotics. It is also recognized to induce gene mutation and promote carcinogenesis. In this review the general concept of nett free radical injury counterbalanced by antioxidants is discussed as oxidative stress. The role of oxidative stress in intestinal ischemia, radiation enteritis, inflammatory bowel disease and the promotion of gastric and colorectal cancer is discussed. The data for the role of oxidative stress in the pathogenesis of ischemic, inflammatory and radiation induced disease are strong, but interventional studies with antioxidants have shown only weak beneficial effects in the above diseases. Therefore the role of antioxidants in the therapy of gastrointestinal diseases remains controversial and should be the subject of controlled trials.

8-iso-prostaglandin F2alpha (8-iso-PGF2alpha), a major F2-isoprostane, is biosynthesized in vivo through nonenzymatic free radical-catalysed peroxidation of arachidonic acid. The levels of F2-isoprostanes both free in the circulation and esterified to the tissue phospholipids increase intensely in animal models of oxidant injury. This study presents the development and validation of a radioimmunoassay of 8-iso-PGF2alpha for the measurement of this substance in the body fluids. Furthermore, its application in normal human volunteers, a pharmacokinetic study performed in rabbits with 8-iso-PGF2alpha, and hepatic lipid peroxidation in rats is reported. An antibody was raised in rabbits by immunization with 8-iso-PGF2alpha coupled to BSA at the carboxylic acid by 1,1'-Carbonyldiimmidazole method. The cross-reactivity of the antibody with 8-iso-15-keto-13,14-dihydro-PGF2alpha, 8-iso-PGF2beta, PGF2alpha, 15-keto-PGF2alpha, 15-keto-13,14-dihydro-PGF2alpha,TXB2, 11beta-PGF2alpha, 9beta-PGF2alpha and 8-iso-PGF3alpha was 1.7, 9.8, 1.1, 0.01, 0.01, 0.1, 0.03, 1.8 and 0.6%, respectively. The intraassay precision was 14.5% (CV) at the level of 64 pg/0.1 ml and 12.2% with 512 pg/0.1 ml in the human plasma. Similarly, intra-assay accuracy was 95.6% and 101% for the low and the high standard, respectively. The detection limit was about 23 pmol/l. The appearance and disappearance of 8-iso-PGF2alpha in the blood and urine following intravenous administration of 8-iso-PGF2alpha in the rabbit was rapid. Free 8-iso-PGF2alpha levels in plasma and urine from normal human volunteers are evaluated and found to correlate with the obtained values by gas chromatography-mass spectrometry methods from other studies. The levels of free 8-iso-PGF2alpha in the plasma and urine increased 7- and 102-fold, respectively, after CCl4 administration to rats. Thus, this 8-iso PGF2alpha radioimmunoassay method is relevant to apply in the oxidative injury studies as an index of in vivo lipid peroxidation through free radical catalysis mechanism.

Since its discovery and isolation the importance of vitamin E in maintaining normal physiologic processes and its value in treating various disease states have been the subject of much controversy. It was our intention to review and highlight some of the arguments and problems regarding the usefulness of vitamin E and to try to put them into proper perspective. The major area of interest concerning vitamin E lies essentially in its role in preventing damage caused by free radicals. The latter are now known to play an important role in radiation induced carcinogenesis, photoaging and photosensitization. The chemistry of vitamin E, its physiological function as a major antioxidant and its interaction with other antioxidants are described by the sum of animal studies, in vitro research and epidemiological investigations. In preparing the current data, it appeared that despite the controversy and conflicting results the body of literature as a whole judges vitamin E to be useful as an antioxidant. Although, in principle, the use of vitamin E can be quite advantageous, the manner of its administration, especially regarding topical application, remains unclear.

Erythromycin (EM) is an antibiotic with potent antiinflammatory effects that is used for treating chronic lower respiratory tract infections. It has been shown that free radicals, such as the superoxide anion and nitric oxide (NO), are pathogenic molecules in viral disease. Much attention has been given to a critical role of NO in the pathologic events of various inflammatory diseases. In the present study, we evaluated the effects of EM on influenza-virus-induced pneumonia in mice infected with a lethal dose of influenza virus A/Kumamoto/Y5/67 (H2N2). The administration of EM at a dose of 3.3 mg/kg/d (intraperitoneally, from Days 1 to 6 after infection), significantly improved the survival rate of mice infected with influenza virus, and the survival rate of the virus-infected mice at Day 20 after infection increased in a dose-dependent fashion with EM administered to the animals, from 14% among controls to 42% among animals given EM at 1.0 mg/kg/d and 57% among those given EM at 3.3 mg/kg/d. The induction of interferon-gamma (IFN-gamma) in the mouse lung was inhibited by EM treatment on Day 6 after infection. Simultaneously, the number of inflammatory cells recovered in lung lavage fluid 6 d after virus infection was significantly reduced by the treatment with EM. The EM treatment resulted in a dose-dependent decrease in the level of nitrite/nitrate (metabolites of NO) in the serum and the NO synthase (NOS)-inducting potential in the lungs of the virus-infected mice. These results indicate that EM may have substantial therapeutic value for various acute inflammatory disorders such as influenza-virus-induced pneumonia, by inhibiting inflammatory-cell responses and suppressing NO overproduction in the lung.

Premarin (Wyeth-Ayerst) is the estrogen replacement treatment of choice and continues to be one of the most widely dispensed prescriptions in the United States. In addition to endogenous estrogens, Premarin contains unsaturated estrogens including equilenin. We synthesized the catechol metabolite of equilenin, 4-hydroxyequilenin (4-OHEN), and found that the semiquinone radical of 4-OHEN reacted with 2'-deoxynucleosides generating very unusual adducts. 2'-Deoxyguanosine (dG), 2'-deoxyadenosine (dA), or 2'-deoxycytosine (dC) all gave four isomers, but no product was observed for thymidine under similar physiological conditions. The structures of these adducts were determined by electrospray mass spectrometry and NMR experiments including 1H, 13C, DQF-COSY, ROESY, HOHAHA, HMQC, and HMBC. The spectral data show that dG forms a cyclic adduct with the 4-OHEN producing 2-N1,3-N2-deoxyguanosyl-1,3-dihydroxy-5,7,9(10)-estratriene-4,17-d ione. Similarly, reaction with dA produced 1-N6,3-C2-deoxyadenosyl-2,3-dihydroxy-5,7,9(10)-estratriene-4,17-d ione, and incubations with dC resulted in 1-N3,3-N4-deoxycytosyl-2,3-dihydroxy-5,7,9(10)-estratriene-4,17-di one. We found that care needed to be taken during the isolation of the dA adducts in particular, as any exposure to acidic environments caused hydrolysis of the sugar moiety leaving alkylated adenine. In mixtures of the deoxynucleosides treated with 4-OHEN, reaction occurred primarily with dG followed by dC and dA. With DNA significant apurinic sites were produced as 4-OHEN-adenine adducts were detected in the ethanol wash prior to hydrolysis. When the DNA was hydrolyzed to deoxynucleosides and analyzed by electrospray mass spectrometry, only one isomer of 4-OHEN-dG and one isomer of 4-OHEN-dC were observed. Our data suggest that several different types of DNA lesions could be expected from 4-OHEN including apurinic sites and bulky stable adducts, in addition to the published oxidized damage to DNA caused by 4-OHEN. The production of these semiquinone radical-derived DNA adducts could play a role in the carcinogenic effects of Premarin estrogens.

Evidence continues to accumulate that oxidative stress is a mediator of endothelial cell dysfunction and thus contributes to the cardiovascular complications of preeclampsia. The mechanisms for the interaction of oxidative stress and endothelial cell function have not been well defined. This review explores potential vasoactive pathways that may be affected by oxidative stress and have been reported to be altered in women with preeclampsia. In pathologic conditions of oxidative stress, increased production of superoxide peroxide anions and nitric oxide has been recognized to inactivate the nitric oxide as a vasorelaxant as well as produce peroxynitrite, a potent oxidant. Increase prostaglandin H (PGH) synthase activity resulting in vasoconstriction predominates in models of oxidative stress. Peroxynitrite increases PGH synthase activity in vitro, providing a potential, but as yet untested, link between oxidative stress, nitric oxide, and PGH synthase pathway, leading to reduced relaxation and increased constriction in the vasculature of women with preeclampsia. Other vasoconstrictors (such as isoprostanes and endothelin) that may be interrelated with oxidative stress and altered endothelial cell function in preeclampsia are also discussed.

Reactive oxygen species derived from molecular oxygen are highly reactive metabolites. These species can be generated by cellular or acellular mechanisms. They react with all biological molecules such as protein, lipid, and carbohydrates. The reaction of these species with lipids, called lipid peroxidation, is a very well-studied phenomenon. Compounds, which scavenge these molecules, are called antioxidants. The disruption of the delicate balance between pro- and antioxidants has been implicated in the pathophysiology of many chronic diseases such as, for example, atherosclerosis. This article presents an introduction to what reactive oxygen species are and their reactions with various metabolites. It deals with lipid peroxidation in detail and with methods for measuring lipid peroxidation. This article also outlines the importance of these species in the pathology of various gynecological diseases.

This review focuses on the question of how oxidative stress in cells populating atherosclerotic lesions stimulates gene expression, cell proliferation, and cell death, and how these events contribute to the initiation, progression, and destablization of the lesions. It is hypothesized that oxidative stress in endothelial cells, macrophages, and smooth muscle cells occurs as a result of the depletion of the cellular content of reduced glutathione. Glutathione becomes oxidized in response to the accumulation of oxidized lipids, the formation of reactive oxygen species released from the mitochondria and generated as part of the activation-induced respiratory burst, and the generation of nitric oxide, peroxynitrite, and thiol radicals. Both in vitro and in vivo evidence suggests that these cells can take up modified lipoproteins that become trapped within the artery wall leading to the overaccumulation of oxidized fatty acids and oxidized forms of cholesterol. The cells also generate oxidized lipids via the activity of lipoxygenases, cyclooxygenases, and myeloperoxidase. A sublethal oxidative stress can activate redox-sensitive kinase cascades and transcription factors such as NFB and AP-1, with resulting increases in the expression of factors associated with an inflammatory response and cellular proliferation. There is also accumulating evidence that suggests that oxidative stress may be associated with the induction of cell death either via stimulation of apoptosis and/or necrosis and that increased cell death contributes to the formation of a necrotic core, the hallmark of an advanced, unstable lesion.

Partial reduction of molecular oxygen can generate reactive oxygen species (ROS), including the hydrogen peroxide, and the free radicals superoxide and hydroxyl. The formation of ROS is a feature of many degenerative diseases, such as atherosclerosis and neurodegeneration, Organisms contain a battery of defense mechanisms to prevent the formation of ROS, to scavenge them, and to repair the damage they cause. Free radicals are also involved in signal transduction pathways. For example, the free radical nitric oxide is involved in signal transduction in both the cardiovascular and central nervous systems. The interplay between nitric oxide and ROS has been a major focus of recent studies, as nitric oxide is an efficient radical scavenger. However, in some cases, such as in the formation of peroxynitrite from nitric oxide and superoxide, the product is potentially more deleterious that the parent radicals. This review describes the major chemical species involved in oxidative stress and free radical biochemistry, and gives a brief overview of their role in pathological conditions.

As women undergo menopause, circulating concentrations of estrogen decrease. The relative estrogen deprivation in postmenopausal women is associated with physiological changes and increased risk of several diseases, including cardiovascular disease. Studies in animals have shown that exogenous estrogen inhibits atherosclerosis, the underlying cause of cardiovascular disease. Ongoing clinical trials will soon provide data for the effect of exogenous estrogen on cardiovascular disease in postmenopausal women. Estrogen has a number of effects that could influence atherogenesis and cardiovascular disease. Estrogens have favorable effects on lipoproteins, but such effects can only account for part of the protection from cardiovascular disease that appears to be conferred by estrogen. Evidence suggests that estrogens can have both prooxidant and antioxidant effects. However, the available evidence suggests that in vivo physiological concentrations of estrogen may have a modest antioxidant activity, and prooxidant activity is unlikely. The antioxidant activity of estrogens and inhibition by estrogens of cellular processes that are thought to promote atherosclerosis are likely to be additional mechanism(s) by which estrogen inhibits atherosclerosis and cardiovascular disease, but more work is needed. Studies of some effects of estrogens on atherogenic processes in isolated cells need to be extended to the whole animal. The influence of estrogen receptors on inhibition of atherosclerosis by estrogen needs to be clarified. Future studies should be designed to investigate separately the estrogenic and antioxidant activities of estrogens and estrogen analogs. Investigations of the antioxidant activities of estrogens should include careful consideration of the interaction of estrogens with endogenous antioxidants and fatty acid saturation, and more attention should be paid to the potential for estrogens to inhibit intraarterial oxidation.

Paroxysmal activity has been reported in the olfactory bulb of rabbits chronically exposed to ozone (O3), postulating that this gas provokes alterations in the electrical activity of the central nervous system. The effects of 1 ppm of O3 inhaled daily for 1 or 3 h prior to each stimulation were studied using the kindling model of epilepsy. Results showed that O3 decreased the duration of the amygdaloid after-discharges during the development of kindling and shortened the latency to the onset of the first generalized seizure in rats exposed for 1 h to O3, while the group exposed for 3 h also decreased the after-discharge duration but delayed the appearance of seizures. Spread of seizure activity, defined as the latency to reach behavioral and electrographic signs of tonic clonic seizures was faster in the 1 h O3 exposed rats and slower in those exposed for 3 h. These findings demonstrate that O3 or its reaction products, such as free radicals, could affect the mechanisms involved in kindling course, such changes being dependent of the time of O3 exposure.

The effect of different doses of methylglyoxal (50-400 mg/kg body wt.) were examined using enzymes involved in the antioxidant function, glutathione (GSH) content and lipid peroxidation in the liver and spleen of Swiss albino mice (7-8 week old) after 6, 12 and 24 h. Significant changes were observed predominantly in the liver. The specific activities of superoxide dismutase (SOD), glutathione-S-transferase (GST), catalase, glyoxalase I (gly I) and glyoxalase II (gly II) were found to decrease in the liver. The mode and magnitude of change in the specific activities was seen to depend on the dose of methylglyoxal and the time after its administration. Methylglyoxal also decreased the GSH content and enhanced the lipid peroxidation in the liver. These findings are suggestive of the adverse effect of methylglyoxal on the antioxidant defence system. It is likely that methylglyoxal undergoes a redox cycle and generates the free radicals which in turn lower the antioxidant status in animals. The increased levels of lipid peroxidation provide support for the involvement of free radical processes in the detrimental effects of methylglyoxal. The response of DT-diaphorase (DTD) seems to be adaptive.

Objectives: Although Alzheimer's disease (AD) is the leading cause of dementia in developed countries, there is an as yet unexplained lower prevalence of the disease in parts of Africa. AD is characterised by a catastrophic loss of neurons; free radicals (oxidative toxins) have been implicated in the destruction of the cells through the process of lipid peroxidative damage of cell membranes. Previously aluminium (Al) and a fragment of beta amyloid (A beta 25-35) were shown to exacerbate free-radical damage, while melatonin reduced this effect. The aim of the present study was: (i) to investigate the conditions determining the toxicity of Al and A beta 25-35; and (ii) to assess whether melatonin could attenuate the damage done by both aluminium and the amyloid fragment, thus suggesting a pathway for the aetiology of AD., Design: An in vitro model system was used in which free radicals were generated, causing lipid peroxidation of platelet membranes, thus simulating the disease process found in the brain., Results: 1. Al and A beta 25-35 caused lipid peroxidation in the presence of the iron (II) ion (Pe2+), Al being more toxic than A beta 25-35. 2. A beta 25-35 attenuated the lipid peroxidation promoted by Al. 3. Hydrogen peroxide (H2O2) greatly exacerbated the toxicity of Al and A beta 25-35. 4. Melatonin prevented lipid peroxidation by Al and A beta 25-35 in the absence of H2O2, but only reduced the process when H2O2 was present., Conclusions: In the light of the results obtained from the present study, the following hypotheses are formulated. 1. In AD, excessive quantities of Al are taken up into the brain, where the Al exacerbates iron-induced lipid peroxidation in the lysosomes. 2. In response, the normal synthetic pathway of amyloid protein is altered to produce A beta fragments which attenuate the toxicity of Al. In the process of sequestering the Al and iron, immature plaques are formed in the brain. 3. Microglia are activated, in an attempt to destroy the plaques by secreting reactive oxygen species such as H2O2. At this point in the disease process, lipid peroxidation causes a catastrophic loss of brain cells. 4. Melatonin, together with other free radical scavengers in the brain, reduces the free-radical damage caused by Al and A beta, except in the latter stages of the disease process. Since melatonin is produced by the pineal gland only in the dark, the excess of electric light in developed countries may help explain why AD is more prevalent in these countries than in rural Africa.

Free radicals are normally produced as a by-product of cellular metabolism. Free radicals are capable of killing bacteria, damage biomolecules, provoke immune response, activate oncogens, cause atherogenesis and enhance ageing process. However, in healthy conditions nature has endowed human body with enormous antioxidant potential. Subtle balance exists between free radical generation and antioxidant defence system to cope with oxidative stress by various enzymes and vitamins at cellular level which prevent the occurrence of disease. However, factors tilting the balance in favour of excess free radicals generation lead to widespread oxidative tissue damage and diseases. Therefore, trouble starts when there is an excess of free radicals and the defence mechanism lags behind. Overwhelming production of free radicals in response to exposure to toxic chemicals and ageing may necessitate judicious antioxidant supplement to help alleviate free radical mediated damage.

Background: The role of monocytes and neutrophils is crucial during acute allograft rejection. They have the capacity to generate toxic reactive oxygen intermediates in response to specific agonists that may act as tissue destructive molecules. We examined the possibility of reactive intermediate-mediated tissue injury in acute lung allograft rejection, as well as the effect of superoxide dismutase., Methods: Allogenic (Brown Norway to F344) or syngenic (F344 to F344) rat left-lung transplantation was performed. Generation of reactive oxygen intermediates in peripheral blood was evaluated by the method of luminol-dependent chemiluminescence. Cell membrane phospholipid peroxidation in the graft was measured as malondialdehyde concentration. The third group of animals having allografts received bovine erythrocyte superoxide dismutase (5,000 U/kg intravenously every 12 hours after transplantation)., Results: Relative chemiluminescence response in the allograft recipient to normal F344 was elevated on postoperative day 1 (257%), then decreased slightly on day 3 (156%) and was elevated again on day 7 (560%) as the process of rejection progressed. Allograft tissue malondialdehyde levels (248.37 +/- 112.35 nM/whole lung, n = 6; p < 0.05 by Student's t test) were higher than isograft levels (139.29 +/- 35.93 nM/whole lung, n = 6) on day 7. Superoxide dismutase treatment significantly ameliorated the histologic degree of rejection on day 7., Conclusions: These results demonstrate the tissue destructive activity of reactive oxygen intermediates during lung allograft rejection. To scavenge free radicals may be a useful therapeutic modality in the management of acute lung allograft rejection.

Colorectal cancer is the second most common cause of cancer deaths in the United States for both sexes. Considerable evidence suggests that the risk of this cancer is increased by the mutagenic actions of free radicals, which are produced during oxidation reactions. Dietary factors, the intestinal flora (bacteria), and endogenously produced metabolites contribute to the production of free radicals in the colon. Dietary antioxidants, such as vitamin E, should reduce the levels of these harmful oxidation products. In the absence of vitamin E, polyunsaturated fats can be oxidized in the colon to produce mutagens, such as lipid hydroperoxides and malondialdehyde. Furthermore, fecal bacteria can generate a high flux of reactive oxygen species (e.g., the superoxide radical [O2*-]) at the surface of the intestinal lumen, and inflammatory cells in close proximity to the colon can produce reactive nitrogen species (e.g., nitrogen dioxide [NO2]). Increasing evidence suggests that the different chemical (e.g., alpha- and gamma-tocopherol) and stereochemical (e.g., RRR- and all-racemic-alpha-tocopherol) forms of vitamin E have distinct biologic potencies, pharmacokinetics, and different abilities to prevent neoplastic transformation. This review considers and evaluates recent studies relating vitamin E and oxidative stress to colon cancer, emphasizing the distinct roles of alpha- and gamma-tocopherols. In addition, recent findings on the antioxidant/pro-oxidant status of the digesta (ingested food) are discussed with respect to the use of antioxidants in chemo-prevention trials for colon cancer.

Oxygen has invaded progressively, and through the ages, an initially anaerobic world. Living organisms had to invent, in the course of evolution, diverse and ingenious defence systems, to survive the toxicity of this element, which was new for them. Strengthened by this experience over billions of years, the present superior organisms, and particularly human species, are thoroughly adapted to 21 per cent of atmospheric oxygen. Nevertheless, the equilibrium is fragile and the menace of oxygen hovers continually. This deleterious potential of oxygen is attributed to the formation, in vivo, of free radicals, a free radical being, by definition, any chemical species possessing one or several mismatched electrons. These free radicals are, in general, very active. They trigger chain reactions able to damage the different constituents of the living organism. Basic oxygen, must be pre-activated to manifest its toxicity. Such an activation can be achieved in two ways: it can be photodynamic, ending mainly in singlet oxygen, it can be reducing, followed by the formation of the anion hydrogen peroxide and of radical hydroxyl; the latter is the most reactive chemical species in the biological world. The reductive process is accelerated in the presence of transition metals, such as iron and copper, and/or specific enzymes (monoxygenase and certain oxydases). This activation takes place in different cellular compartments: mitochondria, microsomes, peroxysomes, cytoplasmic membrane. To this potential toxicity of oxygen the organism opposes different anti-oxidant defence systems. A first group works up the radical chain, inhibiting activation mechanisms. Such a group, as a consequence, warns of the initiation of radical reactions. The second group neutralizes the free radicals already formed and thus stops the chain of propagation. In this group can be found detoxifying enzymes, notably superoxide dismutase and catalase, producing jointly peroxidases, particularly peroxidase glutathions. Such enzymes for the most part have trace elements as cofactors. In this second group can also be found various molecules which act like 'substrate suicide', or as an anti-oxidant shield. Among these molecules, some can act in the lipidic phase, such as tocopherols, carotenoïds and ubiquinones. Other molecules which are lipophobic, mainly ascorbic acid and uric acid, are active in a hydrated environment. In the case of a weakening of such an antioxidant defence or excess production of radicals, a state of oxidative stress occurs. Uncontrolled, these radicals will damage different biological targets: lipids, DNA, proteins. Disturbances of cellular metabolism will occur, unless corrective defences intervene. The identification of these radical phenomena is an obligatory stage. But because of the very short life span of free radicals, identification poses a real analytical problem. However, three approaches are possible: identification of free radicals, either directly by means of paramagnetic electron resonance, or indirectly by identifying some more stable intermediates. evaluation of the traces of radical attack on biological molecules, for example by high performance liquid chromatography, gas-liquid chromatography, colorimetric tests, estimation of the antioxidant status, for example by colorimetric tests, immunoenzymatic methods, high performance liquid chromatography.

To evaluate the age-related response of essential cell functions against peroxidative damage in hyperthermia, we studied the biochemical response to heat stress in both young and aged rats. Passive hyperthermia was immediately observed in rats after exposure to hot environments. In aged rats, the rectal temperature maintained thermal homeostasis and increased to the same degree as in young rats. In these aged animals, the damage from heat stress was more serious than in young animals. In aged rats under normal environmental conditions, hepatic cytosolic glutathione peroxidase (GSH peroxidase) activities were markedly higher than those activities in younger rats. Hepatic cytosolic GSH peroxidase activities were induced by heat stress in young rats but were decreased by hot environments in aged rats. Hepatic catalase activities in young rats were not affected by hot environments, whereas in aged rats, hepatic catalase activities were seriously decreased. Catalase activities in the kidney of aged rats were also reduced by hot environments. Lipid peroxidation in the liver was markedly induced in both young and aged rats. Because the protective enzymes for oxygen radicals in aged rats were decreased by hot environments, lipid peroxidation in the liver was highly induced. In aged rats, lipid peroxidation in intracellular structures such as mitochondria and microsomes was also markedly induced by hot environments. In both young and aged rats, hyperthermia greatly increased the development of hypertrophy and vacuolated degeneration in hepatic cells. In aged rats, both mitochondria and endoplasmic reticulum of the hepatic cells showed serious distortion in shape as a result of exposures to hot environments. Microsomal electron transport systems, such as cytochrome P450 monooxygenase activities, were seriously decreased by heat stress in aged rats but not in young rats. Although the mitochondrial electron transport systems were not affected by acute heat stress in young rats, their activities were simultaneously inhibited after long-lasting heat exposure. In isolated hepatic cells and polymorphonuclear leukocytes in animals, the 70-kDa heat shock-induced proteins were markedly increased by heat stress. In conclusion, the heat stress-inducible oxygen radical damage becomes more severe according to the age of rats. Because aging and hyperthermia have a synergistic effect on lipid peroxidation, protective enzyme activities for oxygen radicals may be essential for surviving and recovering from thermal injury in aged animals and also in humans.

Objectives: Aging processes, and among them brain aging, are thought to be associated with free radical action. It is hypothesized that plasma antioxidant vitamin levels correlate with cognitive performance in healthy older subjects., Design: Longitudinal and cross-sectional comparisons., Setting: The city of Basle, considered representative of the older urban population in Switzerland., Participants: A total of 442 subjects aged 65 to 94 years (mean: 75 years; 312 male, 132 female) was selected from a random sample., Measurements: In 1993, participants were tested for memory, and plasma vitamin levels were measured for the three antioxidants alpha-tocopherol, ascorbic acid, and beta-carotene. These vitamin parameters, measured previously in 1971 in the same sample, were integrated in our analyses. In addition, plasma cholesterol, ferritin, and systolic blood pressure were taken into account. Memory variables were priming, working-memory, free recall, recognition and the WAIS-R vocabulary test (semantic memory)., Results: Correlations showed significant stability of the plasma antioxidants over the time lag of 22 years (alpha-tocopherol: r = .47, P < or = .001; beta-carotene: r = .43, P < .001; ascorbic acid: r = .22, P < .001). Free recall, recognition, and vocabulary (but not priming and working-memory) correlated significantly with ascorbic acid and beta-carotene in the cross-sectional 1993 data as well as in the longitudinal 1971-1993 analysis. These two antioxidants remained significant predictors, especially of semantic memory, after controlling for possible confounding variables like age, education, and gender using multiple regression analyses and ANOVAs., Conclusion: Among people aged 65 and older, higher ascorbic acid and beta-carotene plasma level are associated with better memory performance. These results indicate the important role played by antioxidants in brain aging and may have implications for prevention of progressive cognitive impairments.

Amyotrophic lateral sclerosis (ALS) is a progressive and rapidly fatal neurodegenerative disease in which both upper and lower motoneurones are involved. The recent discovery of mutations affecting the superoxide dismutase (SOD) gene has given impetus to research on the role of oxidative stress in the pathogenesis of familial ALS, while further evidence for a role of excitotoxicity in the disease process has arisen. In this review, Erik Louvel, Jacques Hugon and Adam Doble discuss these findings and, in addition, describe how a number of large, well-controlled clinical trials have taken place to test potential therapies suggested by different aetiological hypotheses, including immunosuppressive therapies, neurotrophic factors, antioxidants and anti-excitotoxic drugs. These trials have led to the first modest steps in the treatment of this devastating neurological disease.

The present study was designed to establish whether or not an increased production of oxygen-derived free radicals is involved in the causation of myringosclerosis. Sclerotic lesions in the tympanic membrane were experimentally elicited by keeping rats with perforated tympanic membranes in an atmosphere containing roughly 40% oxygen. The animals were treated daily with a solution containing either copper zinc-superoxide dismutase plus catalase, deferoxamine, or copper sulfate plus iron chloride, applied to the traumatized area. After 1 week the extension of myringosclerotic plaques was determined otomicroscopically. The pars tensa and pars flaccida were then dissected free and prepared for light microscopic studies. The results showed that treatment with copper zinc-superoxide dismutase plus catalase and deferoxamine inhibited or reduced the development of myringosclerosis, whereas the ears treated with copper sulfate plus iron chloride appeared unaffected. Consequently, the findings support the hypothesis that the formation of oxygen free radicals contributes significantly to the development of myringosclerosis.

The influence of ovarian follicular maturation on lipid peroxidation-induced changes in the fluidity of plasma membrane of granulosa cells was investigated using the fluorogenic polyunsaturated fatty acid cis-parinaric acid (cPNA). An increase in lipid peroxidation results in the decrease of fluorescence intensity of cPNA. A decrease in membrane fluidity results in the decrease of fluorescence polarization of cPNA. Granulosa cells isolated from the largest (F1; mature), and third largest (F3; developing) preovulatory follicles as well as from a pool of immature small yellow follicles (SYF) of the domestic hen ovary were pre-labelled with cPNA. The cPNA-labelled cells were exposed to a free oxygen radical generating agent 2,2'azobis-(2-amidinopropane)-hydrochloride (AAPH). AAPH enhanced the rate of decrease in fluorescence intensity of cPNA-labelled cells. This effect of AAPH was greater in the more differentiated granulosa cells obtained from mature follicles than in less differentiated cells isolated from immature follicles. Similarly, the degree of change in fluorescence polarization was greater in more differentiated F1 granulosa cells than in the less differentiated ones. The analysis of fatty acid composition of phospholipids indicated that the plasma membrane of differentiated granulosa cells contained a higher proportion of the unsaturated fatty acids, oleic and linoleic acids. The fluorescence data show that the rigidity of the plasma membrane of chicken granulosa cells decreases with advancing follicular maturation presumably due to relative increase in the unsaturated fatty acid content of plasma membrane phospholipid.