Effects of piperine on antioxidant pathways in tissues from normal and streptozotocin-induced diabetic rats

Using diabetes mellitus as a model of oxidative damage, this study investigated whether subacute treatment (10 mg/kg/day, intraperitoneally for 14 days) with the compound piperine would protect against diabetes-induced oxidative stress in 30-day streptozotocin-induced diabetic Sprague-Dawley rats. Liver, kidney, brain, and heart were assayed for degree of lipid peroxidation, reduced and oxidized glutathione (GSH and GSSG, respectively) content, and activities of the free-radical detoxifying enzymes catalase, superoxide dismutase, glutathione peroxidase, and glutathione reductase. Piperine treatment of normal rats enhanced hepatic GSSG concentration by 100% and decreased renal GSH concentration by 35% and renal glutathione reductase activity by 25% when compared to normal controls. All tissues from diabetic animals exhibited disturbances in antioxidant defense when compared with normal controls. Treatment with piperine reversed the diabetic effects on GSSG concentration in brain, on renal glutathione peroxidase and superoxide dismutase activities, and on cardiac glutathione reductase activity and lipid peroxidation. Piperine treatment did not reverse the effects of diabetes on hepatic GSH concentrations, lipid peroxidation, or glutathione peroxidase or catalase activities; on renal superoxide dismutase activity; or on cardiac glutathione peroxidase or catalase activities. These data indicate that subacute treatment with piperine for 14 days is only partially effective as an antioxidant therapy in diabetes.     

Exercise conditioning attenuates the hypertensive effects of nitric oxide synthase inhibitor in rat

Many individuals with cardiovascular diseases undergo periodic exercise conditioning with or with out medication. Therefore, this study investigated the interaction of exercise training and chronic nitric oxide synthase (NOS) inhibitor (Nitro-L-Arginine Methyl Ester, L-NAME) treatment on blood pressure and its correlation with aortic nitric oxide (NO), antioxidant defense system and oxidative stress parameters in rats. Fisher 344 rats were divided into four groups: (1) sedentary control, (2) exercise training (ET) for 8 weeks, (3) L-NAME (10 mg/kg, subcutaneous for 8 weeks) and (4) ET + L-NAME. Blood pressure (BP) was monitored weekly for 8 weeks with tail-cuff method. The animals were sacrificed 24 h after last treatments and thoracic aortic rings were isolated and analyzed. Exercise conditioning resulted in a significant increase in respiratory exchange ratio (RER), aortic NO production, NO synthase activity and inducible iNOS protein expression. Training significantly enhanced aortic GSH levels, GSH/GSSG ratio and up-regulation of aortic CuZn-SOD, Mn-SOD, catalase (CAT) glutathione peroxidase (GSH-Px) activity and protein expression and significantly decreased aortic lipid peroxidation. Chronic L-NAME administration resulted in a significant depletion of aortic NO, NOS activity, endothelial (eNOS) and iNOS protein expression, GSH level, GSH/GSSG ratio, down-regulation of aortic antioxidant enzyme activities and protein expressions. Aortic xanthine oxidase (XO) activity significantly increased with increased lipid peroxidation and protein oxidation after L-NAME administration. The biochemical changes were accompanied by increased in BP. Interaction of training and chronic NOS inhibitor treatment resulted in normalization of BP and aortic antioxidant enzyme activity and protein expression, up-regulation of aortic GSH/GSSG ratio, NO levels, Mn-SOD protein expression, depletion of GSSG, protein oxidation and lipid peroxidation. The data suggest that training attenuated the oxidative injury caused by chronic NOS inhibitor treatment by up-regulating the NO and antioxidant systems and lowering the BP in rats.     

Blood glutathione homeostasis as a determinant of resting and exercise-induced oxidative stress in young men

Abstract

Although the importance of glutathione in protection against oxidative stress is well recognised, the role of physiological levels of glutathione and other endogenous antioxidants in protecting against exercise-induced oxidative stress is less clear. We evaluated the role of glutathione and selected antioxidant enzymes as determinants of lipid peroxidation at rest and in response to exercise in men (n = 13–14) aged 20–30 years, who cycled for 40 min at 60% of their maximal oxygen consumption (VO_2max). Levels of plasma thiobarbituric acid reactive substances (plasma TBARS) and blood oxidised glutathione (GSSG) increased by about 50% in response to exercise. Mean blood reduced glutathione (GSH)decreased by 13% with exercise. Of the measured red blood cell (RBC)antioxidant enzyme activities, only selenium-dependent glutathione peroxidase (Se-GPX) activity rose following exercise. In univariate regression analysis, plasma TBARS levels at rest predicted postexercise plasma TBARS and the exercise-induced change in total glutathione (TGSH). Blood GSSG levels at rest were strongly determinant of postexercise levels. Multiple regression analysis showed blood GSH to be a determinant of plasma TBARS at rest. The relative changes in TGSH were determinant of postexercise plasma TBARS. In summary, higher blood GSH and lower plasma TBARS at rest were associated with lower resting, and exercise-induced, lipid peroxidation. Subjects with a favourable blood glutathione redox status at rest maintained a more favourable redox status in response to exercise-induced oxidative stress. Changes in blood GSH and TGSH in response to exercise were closely associated with both resting and exercise-induced plasma lipid peroxidation. These results underscore the critical role of glutathione homeostasis in modulating exercise-induced oxidative stress and, conversely, the effect of oxidative stress at rest on exercise-induced changes in glutathione redox status.     

Adenosine stimulation of fluid transport across rabbit corneal endothelium

Summary The rate of fluid transport across rabbit corneal endothelium has been measured with an automatic volumetric method. The present resolution of the procedure is 1–3 nanoliters, and intervals of measurement can be made as small as seconds. In the presence of glucose, oxidized glutathione (GSSG), and adenosine, the maximal rates were 6.2±1.0 l/hr cm², and 8.2±0.8 l/hr cm² if a large portion of the stroma was dissected away. In the presence of glucose and GSSG only, the rates were lower, namely 3.7±0.5 l/hr cm². The rates consistently increased or decreased when adenosine was added or deleted, respectively, during given experiments. The stimulation of fluid transport by adenosine was in the order of 40–50%. The results raise the possibility that this transport mechanism might be subject to metabolic control.     

Blood glutathione homeostasis as a determinant of resting and exercise-induced oxidative stress in young men

Although the importance of glutathione in protection against oxidative stress is well recognized, the role of physiological levels of glutathione and other endogenous antioxidants in protecting against exercise-induced oxidative stress is less clear. We evaluated the role of glutathione and selected antioxidant enzymes as determinants of lipid peroxidation at rest and in response to exercise in men (n = 13-14) aged 20-30 years, who cycled for 40 min at 60% of their maximal oxygen consumption (VO2max). Levels of plasma thiobarbituric acid reactive substances (plasma TBARS) and blood oxidised glutathione (GSSG) increased by about 50% in response to exercise. Mean blood reduced glutathione (GSH) decreased by 13% with exercise. Of the measured red blood cell (RBC) antioxidant enzyme activities, only selenium-dependent glutathione peroxidase (Se-GPX) activity rose following exercise. In univariate regression analysis, plasma TBARS levels at rest predicted postexercise plasma TBARS and the exercise-induced change in total glutathione (TGSH). Blood GSSG levels at rest were strongly determinant of postexercise levels. Multiple regression analysis showed blood GSH to be a determinant of plasma TBARS at rest. The relative changes in TGSH were determinant of postexercise plasma TBARS. In summary, higher blood GSH and lower plasma TBARS at rest were associated with lower resting, and exercise-induced, lipid peroxidation. Subjects with a favourable blood glutathione redox status at rest maintained a more favourable redox status in response to exercise-induced oxidative stress. Changes in blood GSH and TGSH in response to exercise were closely associated with both resting and exercise-induced plasma lipid peroxidation. These results underscore the critical role of glutathione homeostasis in modulating exercise-induced oxidative stress and, conversely, the effect of oxidative stress at rest on exercise-induced changes in glutathione redox status.     

Action of E. coli endotoxin,IL-1β and TNF-α on antioxidant status of cultured hepatocytes

We have previously reported that endotoxin induces in vivo oxidative stress in liver and a significant increase in hepatic and plasma glutathione concentrations during the acute phase of reversible endotoxic shock in rats. In the present study we examined the in vitro effects of E. coli 0111:B4 endotoxin (lipopolysaccharide, LPS), IL-1 and TNF- on antioxidant status of cultured hepatocytes in order to differentiate between the direct and mediated endotoxin action. LPS increased total glutathione (tGSH) levels after 2 h treatment but decreased oxidized glutathione (GSSG) content which lead to a marked decrease in GSSG/tGSH index. At shorter treatment times a biphasic and dose-dependent behaviour was observed. Cytokines (IL-1 and TNF-) produced significant decreases in tGSH and GSSG after 30 min treatment. Despite its prooxidant effect, TNF- significantly reduced GSSG/tGSH index. Although no significant effects were observed on glutathione reductase activity, both LPS and cytokines induced an important inhibition of glutathione peroxidase which can justify the lipid peroxidation previously observed both in liver during reversible endotoxic shock and in cultured hepatocytes after treatment with endotoxin. The inhibition of hepatic glutathione peroxidase, besides the stimulation of GSH synthesis by LPS and GSH efflux by cytokines, guarantees the export of hepatic glutathione in its reduced form for other organs, contributing to the interorgan homeostasis. On the other hand, the results presented here support a new role for GSSG/tGSH index different from a mere indicator of oxidative stress.     

Lidocaine affects the redox environment and the antioxidant enzymatic system causing oxidative stress in the hippocampus and amygdala of adult rats

AIMS: Our objective was to investigate if oxidative stress is involved in the neural damage caused by lidocaine. MAIN METHODS: Male Wistar rats were used. The control group received 0.9% saline ip and the treated group received a single 60 mg/kg lidocaine dose ip. On days 1, 2, 5, and 10 after dosing, ten rats were sacrificed and their brains were quickly removed. The amygdala and hippocampus were dissected. Five samples were used to determine lipid peroxidation, reactive oxygen species (ROS), reduced glutathione (GSH), and oxidized glutathione (GSSG). Another five were used to measure antioxidant activities of glutathione peroxidase (GPX), catalase, Cu-Zn SOD (superoxide dismutase), Mn SOD, and total SOD. KEY FINDINGS: Ten days after injection of lidocaine, lipid peroxidation increases in the hippocampus because the ROS are enhanced from day 5, whereas in the amygdala lipid peroxidation and the ROS were enhanced only on the first day postinjection. Lidocaine causes an increased concentration of GSH and GSSG in the hippocampus from the first day. In the amygdala the GSH and GSSG content were increased at day 10. In the hippocampus the catalase activity was enhanced, whereas the total SOD and Cu-Zn SOD activities were decreased. In the amygdala the lidocaine enhances the activities of catalase and GPX, but no SOD isoenzymes were modified. SIGNIFICANCE: In this research we demonstrated that lidocaine affects the redox environment and promotes increases of the oxidative markers both in the hippocampus and amygdala but in a different pattern.     

Antioxidant enzyme systems in rat liver and skeletal muscle. Influences of selenium deficiency, chronic training, and acute exercise

The influences of selenium deficiency (Se-D), chronic training, and an acute bout of exercise on hepatic and skeletal muscle antioxidant enzymes, i.e., superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX), as well as glutathione S-transferase (GST) and tissue lipid peroxidation, were investigated in post-weaning male Sprague-Dawley rats. Se-D per se depleted GPX in both liver and skeletal muscle but had no effect on SOD or catalase activity. One hour of treadmill running (20 m/min, 0% grade and 27 m/min, 15% grade for untrained and trained rats, respectively) significantly elevated hepatic catalase and cytosolic SOD activity; more prominent activations were found in the Se-D or untrained rats, whereas skeletal muscle antioxidant enzymes were little affected. Ten weeks of training (1 h/day, 5 days/week at 27 m/min, 15% grade) increased hepatic mitochondrial SOD by 23% (P less than 0.05) in Se-D rats. Both hepatic mitochondrial and cytosolic GPX were decreased by training whereas GPX was increased twofold in skeletal muscle mitochondria. Se-independent GPX was elevated by training only in the skeletal muscle mitochondria of Se-D rats. Lipid peroxidation (malondialdehyde formation) was increased by an acute bout of exercise in hepatic mitochondria of the untrained rats and in skeletal muscle mitochondria of the Se-D rats. These data indicate that antioxidant enzymes in liver and skeletal muscle are capable of adapting to selenium deficiency and exercise to minimize oxidative injury caused by free radicals.     

Ischemia-reperfusion leads to depletion of glutathione content and augmentation of malondialdehyde production in the rat heart from overproduction of oxidants: Can caffeic acid phenethyl ester (CAPE) protect the heart?

During restoration of blood flow of the ischemic heart induced by coronary occlusion, free radicals cause lipid peroxidation with myocardial injury. Lipid peroxidation end-products, such as malondialdehyde (MDA), have been used to assess oxygen free radical-mediated injury of the ischemic-reperfused (I/R) myocardium in rats. This experimental study assessed the preventive effect of caffeic acid phenthyl ester (CAPE), antioxidant, on I/R-induced lipid peroxidation in the rat heart. We are also interested in the role of CAPE on glutathione (GSH) levels, an antioxidant whose levels are influenced by oxidative stress. I/R leads to the depletion of GSH which is the major intracellular nonprotein sulphydryl and plays an important role in the maintenance of cellular proteins and lipid in their functional state and acts primarily to protect these important structures against the threat of oxidation. In addition, we also examined morphologic changes in the heart by using light microscopy. The left coronary artery was occluded for 30 min and then reperfused for 120 min more before the experiment was terminated. CAPE (50 M kg^–1) was administered 10 min prior to ischemia and during occlusion by infusion. At the end of the reperfusion period, rats were sacrificed, and the heart was quickly removed for biochemical determination and histopathological analysis. I/R was accompanied by a significant increase in MDA production and decrease in GSH content in the rat heart. Administration of CAPE reduced MDA production and prevented depletion of GSH content. These beneficial changes in these biochemical parameters were also associated with parallel changes in histopathological appearance. These findings imply that I/R plays a causal role in heart injury due to overproduction of oxygen radicals or insufficient antioxidant and CAPE exert cardioprotective effects probably by the radical scavenging and antioxidant activities.     

Taurine restores ethanol-induced depletion of antioxidants and attenuates oxidative stress in rat tissues

Summary. Ethanol by its property of generating free radicals during the course of its metabolism causes damage to cell structure and function. The study investigates the protective effects of the antioxidant aminoacid taurine on ethanol-induced lipid peroxidation and antioxidant status. Male Wistar rats of body weight 170–190g were divided into 4 groups and maintained for 28 days as follows: a control group and taurine-supplemented control group, taurine supplemented and unsupplemented ethanol-fed group. Ethanol was administered to rats at a dosage of 3g/kg body weight twice daily and taurine was provided in the diet (10g/kg diet). Lipid peroxidation products and antioxidant potential were quantitated in plasma and in following tissues liver, brain, kidney and heart.Increased levels of thiobarbituric acid substances (TBARS) and lipid hydroperoxides (LHP) in plasma and tissues, decreased activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) were observed in hemolysate and tissues of ethanol-fed rats. The contents of reduced glutathione (GSH), -tocopherol and ascorbic acid in plasma and tissues were significantly reduced in these animals as compared to control animals. Simultaneous administration of taurine along with ethanol attenuated the lipid peroxidation process and restored the levels of enzymatic and non-enzymatic antioxidants. We propose that taurine may have a bioprotective effect on ethanol-induced oxidative stress.     

Effects of new antioxidant compounds PNU-104067F and PNU-74389G on antioxidant defense in normal and diabetic rats

Diabetes mellitus and its complications are associated with elevated oxidative stress, leading to much interest in antioxidant compounds as possible therapeutic agents. Two new classes of antioxidant compounds, the pyrrolopyrimidines and the 21-aminosteroids, are known to inhibit lipid peroxidation and other biomolecular oxidation. We hypothesized that in the presence of excess oxidants or the impaired antioxidant defense seen in diabetes mellitus, administration of antioxidants such as these may reverse the effects of diabetes on antioxidant parameters. This study measured the effects of subchronic (14 day) treatment with a pyrrolopyrimidine (PNU-104067F) or a 21-aminosteroid (PNU-74389G) in normal and diabetic Sprague-Dawley rats. Activity levels of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, concentrations of oxidized and reduced glutathione, and lipid peroxidation were used as measures of antioxidant defense in liver, kidney, heart, and brain tissue. In normal rats, the only effect was a 43% increase in cardiac lipid peroxidation after treatment with PNU-104067F. In diabetic rats, the only reversals of the effects of diabetes were a 30% decrease in hepatic glutathione peroxidase activity after PNU-74389G treatment and a 33% increase in cardiac glutathione disulfide concentration after PNU-104067F treatment. In contrast to these effects, increased cardiac glutathione peroxidase and catalase activities, increased brain glutathione peroxidase activity, increased hepatic lipid peroxidation, decreased hepatic glutathione content, and decreased hepatic catalase activity were seen in diabetic rats, reflecting an exacerbation of the effects of diabetes.     

The effect of adenosine and adenosine analogues on methylxanthine-induced hypercalciuria in the rat

The chronic effects of dietary caffeine, theophylline, and theobromine on urinary calcium excretion were investigated in the adult male rat. The effect of acutely administered adenosine and adenosine analogues on methylxanthine-induced hypercalciuria was concurrently investigated. When rats were fed equimolar amounts of theobromine, caffeine, and theophylline, it was found that urinary calcium excretion increased; on day 7 values were increased over controls (p less than 0.05) by 54, 146, and 208%, respectively. On day 20, an injection of adenosine reduced calcium excretion in methylxanthine-treated rats to levels not different from control values. In another series of experiments, theophylline was fed to a group of rats to establish hypercalciuria. Three adenosine analogues N6-(L-2-phenylisopropyl)adenosine (R-PIA), N6-(D-2-phenylisopropyl)adenosine (S-PIA), and 5'-(N-ethylcarboxamido)adenosine (NECA) with different adenosine receptor specificities (A2,A1, and weakly A2, respectively) were administered to different groups of theophylline-fed and control rats, and their calcium excretions were measured. It was found that the order of efficacy of the analogues in reducing calcium excretion was NECA greater than R-PIA greater than S-PIA, which is consistent with the receptor being A2. A proportion of the methylxanthine-induced hypercalciuria may be due to adenosine antagonism.     

Effects of endurance training and acute exhaustive exercise on antioxidant defense mechanisms in rat heart

We investigated whether 8-week treadmill training strengthens antioxidant enzymes and decreases lipid peroxidation in rat heart. The effects of acute exhaustive exercise were also investigated. Male rats (Rattus norvegicus, Sprague-Dawley strain) were divided into trained and untrained groups. Both groups were further divided equally into two groups where the rats were studied at rest and immediately after exhaustive exercise. Endurance training consisted of treadmill running 1.5 h day(-1), 5 days week(-1) for 8 weeks. For acute exhaustive exercise, graded treadmill running was conducted. Malondialdehyde level in heart tissue was not affected by acute exhaustive exercise in untrained and trained rats. The activities of glutathione peroxidase and glutathione reductase enzymes decreased by both acute exercise and training. Glutathione S-transferase and catalase activities were not affected. Total and non-enzymatic superoxide scavenger activities were not affected either. Superoxide dismutase activity decreased by acute exercise in untrained rats; however, this decrease was not observed in trained rats. Our results suggested that rat heart has sufficient antioxidant enzyme capacity to cope with exercise-induced oxidative stress, and adaptive changes in antioxidant enzymes due to endurance training are limited.     

Modulation of cyclophosphamide-induced early lung injury by curcumin,an anti-inflammatory antioxidant

Cyclophosphamide causes lung injury in rats through its ability to generate free radicals with subsequent endothelial and epithelial cell damage. In order to observe the protective effects of a potent anti-inflammatory antioxidant, curcumin (diferuloyl methane) on cyclophosphamide-induced early lung injury, healthy pathogen free male Wistar rats were exposed to 20 mg/100 g body weight of cyclophosphamide, intraperitoneally as a single injection. Prior to cyclophosphamide intoxication oral administration of curcumin was performed daily for 7 days. At various time intervals (2, 3, 5 and 7 days post insult) serum and lung samples were analyzed for angiotensin converting enzyme, lipid peroxidation, reduced glutathione and ascorbic acid. Bronchoalveolar lavage fluid was analyzed for biochemical constituents. The lavage cells were examined for lipid peroxidation and glutathione content. Excised lungs were analyzed for antioxidant enzyme levels. Biochemical analyses revealed time course increases in lavage fluid total protein, albumin, angiotensin converting enzyme (ACE), lactate dehydrogenase, N-acetyl--D-glucosaminidase, alkaline phosphatase, acid phosphatase, lipid peroxide levels and decreased levels of glutathione (GSH) and ascorbic acid 2, 3, 5 and 7 days after cyclophosphamide intoxication. Increased levels of lipid peroxidation and decreased levels of glutathione and ascorbic acid were seen in serum, lung tissue and lavage cells of cyclophosphamide groups. Serum angiotensin converting enzyme activity increased which coincided with the decrease in lung tissue levels. Activities of antioxidant enzymes were reduced with time in the lungs of cyclophosphamide groups. However, a significant reduction in lavage fluid biochemical constituents, lipid peroxidation products in serum, lung and lavage cells with concomitant increase in antioxidant defense mechanisms occurred in curcumin fed cyclophosphamide rats. Therefore, our results suggest that curcumin is effective in moderating the cyclophosphamide induced early lung injury and the oxidant-antioxidant imbalance was partly abolished by restoring the glutathione (GSH) with decreased levels of lipid peroxidation.     

Effects of aging on the susceptibility to the toxic effects of cyclosporin A in rats. Changes in liver glutathione and antioxidant enzymes

Free radicals are involved in aging and cyclosporin A-induced toxicity. The age-related changes in the liver oxidative status of glutathione, lipid peroxidation, and the activity of the enzymatic antioxidant defense system, as well as the influence of aging on the susceptibility to the hepatotoxic effects of cyclosporin (CyA) were investigated in rats of different ages (1, 2, 4, and 24 months). The hepatic content of reduced glutathione (GSH) increased with aging, peaked at 4 months, and decreased in senescent rats. By contrast, glutathione disulfide (GSSG) and thiobarbituric acid-reactive substances (TBARS) concentrations and superoxide dismutase, catalase, and glutathione peroxidase activities were higher in the oldest than in the youngest rats. CyA treatment, besides inducing the well-known cholestatic syndrome, increased liver GSSG and TBARS contents and the GSSG/GSH molar ratio, and altered the nonenzymatic and enzymatic antioxidant defense systems. The CyA-induced cholestasis and hepatic depletion of GSH, and the increases in the GSSG/GSH ratio, and in GSSG and TBARS concentrations were higher in the older than the mature rats. Moreover, superoxide dismutase and catalase activities were found to be significantly decreased only in treated senescent rats. The higher CyA-induced oxidative stress, lipoperoxidation, and decreases in the antioxidant defense systems in the aged animals render them more susceptible to the hepatotoxic effects of cyclosporin.     

Antagonism by theophylline of the adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine at the guinea pig ileum

The inhibitory effect of the putative adenosine A2 receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA) on acetylcholine release from the stimulated guinea pig ileum preparation and the nature of its antagonism by theophylline were investigated. NECA was shown to inhibit the response of the ileum preparation in a dose-dependent fashion, and an EC50 value of 1.62 X 10(-8) M was determined. This value was comparable with that determined for the A1 receptor agonist N6-R-phenylisopropyladenosine (R-PIA) (2.57 X 10(-8) M) using the same preparation. Competitive antagonism of the inhibitory effect of NECA by theophylline was quantitated and a pA2 value of 5.04 for the methylxanthine was obtained. This value was similar to those obtained previously for R-PIA and adenosine itself and suggests that these nucleosides may be interacting with the same receptor site on myenteric nerve endings. These findings do not permit the designation of the receptor as an A1 or A2 subtype according to current criteria.     

Aging and acute exercise enhance free radical generation in rat skeletal muscle.

Reactive oxygen species (ROS) are implicated in the mechanism of biological aging and exercise-induced oxidative damage. The present study examined the effect of an acute bout of exercise on intracellular ROS production, lipid and protein peroxidation, and GSH status in the skeletal muscle of young adult (8 mo, n = 24) and old (24 mo, n = 24) female Fischer 344 rats. Young rats ran on a treadmill at 25 m/min and 5% grade until exhaustion (55.4 +/- 2.7 min), whereas old rats ran at 15 m/min and 5% grade until exhaustion (58.0 +/- 2.7 min). Rate of dichlorofluorescin (DCFH) oxidation, an indication of ROS and other intracellular oxidants production in the homogenate of deep vastus lateralis, was 77% (P < 0.01) higher in rested old vs. young rats. Exercise increased DCFH oxidation by 38% (P < 0.09) and 50% (P < 0.01) in the young and old rats, respectively. DCFH oxidation in isolated deep vastus lateralis mitochondria with site 1 substrates was elevated by 57% (P < 0.01) in old vs. young rats but was unaltered with exercise. Significantly higher DCFH oxidation rate was also found in aged-muscle mitochondria (P < 0.01), but not in homogenates, when ADP, NADPH, and Fe(3+) were included in the assay medium without substrates. Lipid peroxidation in muscle measured by malondialdehyde content showed no age effect, but was increased by 20% (P < 0.05) with exercise in both young and old rats. Muscle protein carbonyl formation was unaffected by either age or exercise. Mitochondrial GSH/ GSSG ratio was significantly higher in aged vs. young rats (P < 0.05), whereas exercise increased GSSG content and decreased GSH/GSSG in both age groups (P < 0.05). These data provided direct evidence that oxidant production in skeletal muscle is increased in old age and during prolonged exercise, with both mitochondrial respiratory chain and NADPH oxidase as potential sources. The alterations of muscle lipid peroxidation and mitochondrial GSH status were consistent with these conclusions.     

Anti-oxidant effects of cinnamon (Cinnamomum verum) bark and greater cardamom (Amomum subulatum) seeds in rats fed high fat diet

In order to gain insight into the antioxidant effect of cinnamon (Cinnamomum verum; Lauraceae) and cardamom (Amomum subulatum; Zingiberaceae) hepatic and cardiac antioxidant enzymes, glutathione (GSH) content and lipid conjugated dienes were studied in rats fed high fat diet along with cinnamon or cardamom. The antioxidant enzyme activities were found to be significantly enhanced whereas GSH content was markedly restored in rats fed a fat diet with spices. In addition, these spices partially counteracted increase in lipid conjugated dienes and hydroperoxides, the primary products of lipid peroxidation. Thus, it appears that these spices exert antioxidant protection through their ability to activate the antioxidant enzymes.     

Selenium deficiency potentiates paraquat-induced lipid peroxidation in isolated perfused rat lung

Glutathione peroxidase (GSHPx), a seleno-enzyme, reduces lipid hydroperoxides while producing oxidized glutathione (GSSG), which can efflux from cells. To study the role of GSHPx in antioxidant defense, isolated lungs from selenium-deficient rats were perfused for 2 h with or without 1 mM paraquat. Perfusate GSSG was measured as an index of GSHPx activity, and malondialdehyde (MDA) as an index of lipid peroxidation. Selenium deficiency decreased lung GSHPx activity 75-80%. During perfusion control lungs showed GSSG efflux of 8.5 +/- 4.5 nmol/h and with paraquat 49.1 +/- 12.1 nmol/h. Selenium-deficient lungs with or without paraquat showed GSSG efflux of 16.4 +/- 5.3 and 13.7 +/- 8.9 nmol/h, respectively. MDA efflux occurred only in paraquat-perfused selenium-deficient lungs (7.8 +/- 2.7 nmol/h). Lung homogenates from this group had lower GSH + GSSG than the other three groups. These results indicate an inverse correlation between GSSG efflux and MDA accumulation from paraquat-perfused lungs and suggest that increased turnover of the GSHPx reaction protects paraquat-perfused lungs from lipid peroxidation.