Glucose-6-phosphate dehydrogenase activity and NADPH/NADP+ ratio in liver and pancreas are dependent on the severity of hyperglycemia in rat

Hyperglycemia is associated with metabolic disturbances affecting cell redox potential, particularly the NADPH/NADP+ ratio and reduced glutathione levels. Under oxidative stress, the NADPH supply for reduced glutathione regeneration is dependent on glucose-6-phosphate dehydrogenase. We assessed the effect of different hyperglycemic conditions on enzymatic activities involved in glutathione regeneration (glucose-6-phosphate dehydrogenase and glutathione reductase), NADP(H) and reduced glutathione concentrations in order to analyze the relative role of these enzymes in the control of glutathione restoration. Male Sprague-Dawley rats with mild, moderate and severe hyperglycemia were obtained using different regimens of streptozotocin and nicotinamide. Fifteen days after treatment, rats were killed and enzymatic activities, NADP(H) and reduced glutathione were measured in liver and pancreas. Severe hyperglycemia was associated with decreased body weight, plasma insulin, glucose-6-phosphate dehydrogenase activity, NADPH/NADP+ ratio and glutathione levels in the liver and pancreas, and enhanced NADP+ and glutathione reductase activity in the liver. Moderate hyperglycemia caused similar changes, although body weight and liver NADP+ concentration were not affected and pancreatic glutathione reductase activity decreased. Mild hyperglycemia was associated with a reduction in pancreatic glucose-6-phosphate dehydrogenase activity. Glucose-6-phosphate dehydrogenase, NADPH/NADP+ ratio and glutathione level, vary inversely in relation to blood glucose concentrations, whereas liver glutathione reductase was enhanced during severe hyperglycemia. We conclude that glucose-6-phosphate dehydrogenase and NADPH/NADP+ were highly sensitive to low levels of hyperglycemia. NADPH/NADP+ is regulated by glucose-6-phosphate dehydrogenase in the liver and pancreas, whereas levels of reduced glutathione are mainly dependent on the NADPH supply.     

Oxidant/antioxidant status in blood of patients with malignant breast tumour and benign breast disease

The aim of this study was to investigate the alterations in lipid peroxidation and antioxidant enzyme defences in the blood of patients with malignant breast tumour and benign breast disease. Forty patients with malignant breast tumour, 20 patients with benign breast disease and also 20 healthy control subjects were recruited for the study. Malondialdehyde levels in plasma and erythrocytes, and the activities of erythrocyte CuZn-superoxide dismutase, catalase, glutathione peroxidase and glucose-6-phosphate dehydrogenase were measured. Malondialdehyde levels were higher in patients with both benign breast disease and malignant breast tumour compared with control subjects. The activities of all antioxidant enzymes were higher in patients with malignant breast tumour, while only glutathione peroxidase and CuZn-superoxide dismutase activities were higher in patients with benign breast disease. Except for glucose-6-phosphate dehydrogenase, the antioxidant enzymes studied correlated positively with the malondialdehyde levels in patients with malignant breast tumour. On the other hand, only glucose-6-phosphate dehydrogenase activity was increased by the level of malignancy. The activity increases in erythrocyte antioxidant enzymes may be a compensatory upregulation in response to increased oxidative stress especially in patients with malignant breast tumour.     

Oxidative stress reversibly inactivates myocardial enzymes during cardiac arrest

Oxidative stress during cardiac arrest may inactivate myocardial enzymes and thereby exacerbate ischemic derangements of myocardial metabolism. This study examined the impact of cardiac arrest on left ventricular enzymes. Beagles were subjected to 5 min of cardiac arrest and 5 min of open-chest cardiac compressions (OCCC) before epicardial direct current countershocks were applied to restore sinus rhythm. Glutathione/glutathione disulfide redox state (GSH/GSSG) and a panel of enzyme activities were measured in snap-frozen left ventricle. To test whether oxidative stress during arrest inactivated the enzymes, metabolic (pyruvate) or pharmacological (N-acetyl-l-cysteine) antioxidants were infused intravenously for 30 min before arrest. During cardiac arrest, activities of phosphofructokinase, citrate synthase, aconitase, malate dehydrogenase, creatine kinase, glucose-6-phosphate dehydrogenase, and glutathione reductase fell by 56, 81, 55, 34, 42, 55, and 45%, respectively, coincident with 50% decline in GSH/GSSG. OCCC effected full recovery of glutathione reductase and partial recovery of citrate synthase and aconitase, in parallel with GSH/GSSG. Phosphofructokinase, malate dehydrogenase, creatine kinase, and glucose-6-phosphate dehydrogenase recovered only after cardioversion. Antioxidant pretreatments augmented phosphofructokinase, aconitase, and malate dehydrogenase activities before arrest and enhanced these activities, as well as those of citrate synthase and glucose-6-phosphate dehydrogenase, during arrest. In conclusion, cardiac arrest reversibly inactivates several important myocardial metabolic enzymes. Antioxidant protection of these enzymes implicates oxidative stress as a principal mechanism of enzyme inactivation during arrest.     

Oxidant-antioxidant imbalance in the erythrocytes of sporadic amyotrophic lateral sclerosis patients correlates with the progression of disease

Free radicals are implicated in numerous disease processes including motor neuron degeneration (MND). Antioxidant defense enzymes: superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSHPx), glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G-6-PDH) in the erythrocytes are capable of detoxifying reactive oxygen species produced endogenously or exogenously. In the present study, the extent of lipid peroxidation (LPO) and antioxidant defenses were evaluated in the erythrocytes of 20 sporadic amyotrophic lateral sclerosis (ALS) patients and 20 controls. We observed that lipid peroxidation in the erythrocytes of amyotrophic lateral sclerosis patients significantly increased with respect to controls (P<0.001). On the other hand, catalase activity was found to be significantly lower (P<0.001). The activities of glucose-6-phosphate dehydrogenase, glutathione reductase and glutathione levels were also found to be significantly reduced in ALS patients compared to healthy subjects (P<0.001, P<0.01 and P<0.01, respectively). It was further observed that lipid peroxidation started to increase and catalase, glutathione reductase, glucose-6-phosphate dehydrogenase enzyme activities and glutathione levels started to decrease as amyotrophic lateral sclerosis progressed from 6 to 24 months, suggesting a correlation between these parameters and duration of amyotrophic lateral sclerosis. This study confirms the involvement of oxidative stress during the progression of amyotrophic lateral sclerosis and the need to develop specific peripheral biomarkers.     

Phenylpyruvic Acid decreases glucose-6-phosphate dehydrogenase activity in rat brain

Phenylketonuria is a recessive autosomal disorder that is caused by a deficiency in the activity of phenylalanine-4-hydroxylase, which converts phenylalanine to tyrosine, leading to the accumulation of phenylalanine and its metabolites phenyllactic acid, phenylacetic acid, and phenylpyruvic acid in the blood and tissues of patients. Phenylketonuria is characterized by severe neurological symptoms, but the mechanisms underlying brain damage have not been clarified. Recent studies have shown the involvement of oxidative stress in the neuropathology of hyperphenylalaninemia. Glucose-6-phosphate dehydrogenase plays an important role in antioxidant defense because it is the main source of reduced nicotinamide adenine dinucleotide phosphate (NADPH), providing a reducing power that is essential in protecting cells against oxidative stress. Therefore, the present study investigated the in vitro effect of phenylalanine (0.5, 1, 2.5, and 5?mM) and its metabolites phenyllactic acid, phenylacetic acid, and phenylpyruvic acid (0.2, 0.6, and 1.2?mM) on the activity of enzymes of the pentose phosphate pathway, which is involved in the oxidative phase in rat brain homogenates. 6-Phosphogluconate dehydrogenase activity was not altered by any of the substances tested. Phenylalanine, phenyllactic acid, and phenylacetic acid had no effect on glucose-6-phosphate dehydrogenase activity. Phenylpyruvic acid significantly reduced glucose-6-phosphate dehydrogenase activity without pre-incubation and after 1?h of pre-incubation with the homogenates. The inhibition of glucose-6-phosphate dehydrogenase activity caused by phenylpyruvic acid could elicit an impairment of NADPH production and might eventually alter the cellular redox status. The role of phenylpyruvic acid in the pathophysiological mechanisms of phenylketonuria remains unknown.     

Ethanol-induced alterations of the antioxidant defense system in rat kidney

We report here the effects of chronic ethanol consumption on the antioxidant defense system in rat kidney. Thirty-two male Wistar rats were randomly divided in two identical groups and were treated as follows: control group (water for fluid) and the ethanol-fed group (2 g/kg body weight/24 h). The animals were sacrificed after 10 weeks, and respectively 30 weeks of ethanol consumption, and the renal tissue was isolated and analyzed. Results revealed that kidney alcohol dehydrogenase activities increased significantly after ethanol administration, but the electrophoretic pattern of alcohol dehydrogenase isoforms was unmodified. The SDS polyacrylamidegel electrophoretic study of kidney proteins has revealed the appearance of two new protein bands after long-term ethanol consumption. The kidney reduced glutathione/oxidized glutathione ratio decreased, indicating an oxidative stress response due to ethanol ingestion. The malondialdehyde contents and xanthine oxidase activities were unchanged. The antioxidant enzymatic defense system showed a different response during the two periods of ethanol administration. After 10 weeks, catalase, glutathione peroxidase, glutathione reductase, and glucose-6-phosphate dehydrogenase were activated, while superoxide dismutase, glutathione transferase, and gamma-glutamyltranspeptidase levels were stationary. After 30 weeks, superoxide dismutase and glutathione peroxidase activities were unmodified, but catalase, glutathione transferase, gamma-glutamyltranspeptidase, glutathione reductase, and glucose-6-phosphate dehydrogenase activities were significantly increased. Remarkable changes have been registered after 30 weeks of ethanol administration for glutathione reductase and glucose-6-phosphate dehydrogenase activities, including an increase by 106 and 216' of control values, respectively. These results showed specific changes in rat kidney antioxidant system and glutathione status as a consequence of long-term ethanol administration.     

The dehydrogenase-mediated recycling of NADPH is a key antioxidant system against salt-induced oxidative stress in olive plants

NADPH is an important molecule in the redox balance of the cell. In this paper, using olive tissue cultures as a model of the function of the NADPH-generating dehydrogenases in the mechanism of oxidative stress induced by severe salinity conditions was studied. When olive (Olea europaea) plants were grown with 200 mM NaCl, a 40% reduction in leaf fresh weight was produced. The content of non-enzymatic antioxidants such as ascorbate and glutathione was diminished between 20% to 39%, whereas the H2O2 content was increased threefold. In contrast, the analysis of the activity and protein contents of the main antioxidative enzymes showed a significant increase of catalase, superoxide dismutase and glutathione reductase. Overall, these changes strongly suggests that NaCl induces oxidative stress in olive plants. On the other hand, while the content of glucose-6-phosphate was increased almost eightfold in leaves of plants grown under salt stress, the content of NAD(P)H (reduced and oxided forms) did not show significant variations. Under salt stress conditions, the activity and protein contents of the main NADPH-recycling enzymes, glucose-6-phosphate dehydrogenase (G6PDH), isocitrate dehydrogenase (ICDH), malic enzyme (ME) and ferrodoxin-NADP reductase (FNR) showed an enhancement of 30-50%. In leaves of olive plants grown with 200 mM NaCl, analysis of G6PDH by immunocytochemistry and confocal laser scanning microscopy showed a general increase of this protein in epidermis, palisade and spongy mesophyll cells. These results indicate that in olive plants, salinity causes reactive oxygen species (ROS)-mediated oxidative stress, and plants respond to this situation by inducing different antioxidative enzymes, especially the NADPH-producing dehydrogenases in order to recycle NADPH necessary for the protection against oxidative damages. These NADP-dehydrogenases appear to be key antioxidative enzymes in olive plants under salt stress conditions.     

Blood glutathione redox status in gestational hypertension

Gestational hypertension during the third trimester reflects an exaggerated maternal inflammatory response to pregnancy. We hypothesized that oxidative stress present even in normal pregnancy becomes uncompensated in hypertensive patients. A glucose-6-phosphate dehydrogenase (G6PD) activity sufficient to meet the increased reductive equivalent need of the cells is indispensable for defense against oxidative stress. The erythrocyte glutathione redox system was studied, where G6PD is the only NADPH source. The glutathione (GSH) redox status was measured both in vivo and after an in vitro oxidative challenge in pregnant women with gestational hypertension (n = 19) vs. normotensive pregnant subjects (n = 18) and controls (n = 20). An erythrocyte GSH depletion with an increase in the oxidized form (GSSG) resulted in an elevated ratio GSSG/GSH (0.305 +/- 0.057; mean +/- SD) in hypertensive pregnant women vs. normotensive pregnant or control subjects (0.154 +/- 0.025; 0.168 +/- 0.073; p <.001). In hypertensive pregnant patients, a "GSH stability" decrease after an in vitro oxidative challenge suggested a reduced GSH recycling capacity resulting from an insufficient NADPH supply. The erythrocyte GSSG/GSH ratio may serve as an early and sensitive parameter of the oxidative imbalance and a relevant target for future clinical trials to control the effects of antioxidant treatment in women at increased risk of the pre-eclampsia syndrome.     

Mathematical modelling of metabolic pathways affected by an enzyme deficiency. Energy and redox metabolism of glucose-6-phosphate-dehydrogenase-deficient erythrocytes

The effects of various forms of glucose-6-phosphate dehydrogenase deficiency on erythrocyte metabolism have been studied on the basis of a complex mathematical model which comprises the main pathways of this cell: glycolysis, pentose pathway, reactions of the glutathione and adenine nucleotide metabolism. The calculated flux rates through the oxidative pentose pathway with and without methylene blue are in good accord with experimental results. The degree of deficiency as predicted by the model on the basis of calculated upper oxidative load boundaries, as well as of maximal methylene blue stimulation, correlates with the individual clinical manifestation of the metabolic disease. Therefore, the model allows one to judge the degree of metabolic disorder in the presence of glucose-6-phosphate dehydrogenase enzymopathies if the kinetic properties of the defect enzyme are known. Experimentally accessible parameters for an assessment of the oxidative load capacity of cells in vivo are proposed. It is pointed out that the threshold of tolerance as to energetic load is drastically reduced in the case of severe glucose-6-phosphate dehydrogenase deficiency.     

NADP+ and NADPH in glucose-6-phosphate dehydrogenase-deficient erythrocytes under oxidative stimulation.

A mild oxidative stimulation of the hexose monophosphate pathway of human glucose-6-phosphate dehydrogenase (EC 1.1.1.49)-deficient erythrocytes (Mediterranean variant) causes a significant drop in NADPH. These results, other than to confirm that glucose-6-phosphate dehydrogenase deficiency is a product deficiency disorder, demonstrate that under oxidative stimulation glutathione reductase may become functionally impaired and GSSG cannot be reduced at a sufficient rate.     

Mammalian small stress proteins protect against oxidative stress through their ability to increase glucose-6-phosphate dehydrogenase activity and by maintaining optimal cellular detoxifying machinery

The protective activity of small stress proteins (sHsp) against H2O2-mediated cell death in the highly sensitive murine L929 fibroblast has been analyzed. We report here that the human Hsp27- and murine Hsp25-mediated rise in glutathione (GSH) levels as well as the maintenance of this redox modulator in its reduced form was directly responsible for the protection observed at the level of cell morphology and mitochondrial membrane potential. sHsp expression also buffered the increase in protein oxidation following H2O2 treatment and protected several key enzymes against inactivation. In this case, however, the protection necessitated both an increase in GSH and the presence of sHsp per se since the pattern of protection against protein oxidation mediated by a simple GSH increase was different from that induced by sHsp expression. Among the enzymes analyzed, we noticed that sHsp significantly increased glucose-6-phosphate dehydrogenase (G6PD) activity and to a lesser extent glutathione reductase and glutathione transferase activities. Moreover, an increased GSH level was observed in G6PD-overexpressing L929 cell clones. Taken together our results suggest that sHsp protect against oxidative stress through a G6PD-dependent ability to increase and uphold GSH in its reduced form and by using this redox modulator as an essential parameter of their in vivo chaperone activity against oxidized proteins.     

NADP and NADPH in glucose-6-phosphate dehydrogenase-deficient erythrocytes under oxidative stimulation

A mild oxidative stimulation of the hexose monophosphate pathway of human glucose-6-phosphate dehydrogenase (EC 1.1.1.49)-deficient erythrocytes (Mediterranean variant) causes a significant drop in NADPH.

These results, other than to confirm that glucose-6-phosphate dehydrogenase deficiency is a product deficiency disorder, demonstrate that under oxidative stimulation glutathione reductase may become functionally impaired and GSSG cannot be reduced at a sufficient rate.     

Acephate induced oxidative damage in erythrocytes

The effect of oral administration of acephate (360 mg/kg body weight), for 15 days, daily, was investigated on the erythrocytes of male rats. Activities of acetyl cholinesterase and glucose-6-phosphate dehydrogenase decreased, while those of glutathione-s-transferase and glutathione reductase increased. Decreased glutathione content and increased lipid peroxidation suggest that there was increased oxidative stress in the erythrocytes of treated animals. Increased cholesterol/phospholipid ratio in the erythrocyte membranes and morphological changes in RBCs (scanning electron microscopy studies) were observed in acephate treated animals. The results clearly suggest that acephate induced oxidative stress in erythrocytes leads to morphological changes.     

Hyperoxia results in transient oxidative stress and an adaptive response by antioxidant enzymes in goldfish tissues

The effects of hyperoxia on the status of antioxidant defenses and markers of oxidative damage were evaluated in goldfish tissues. The levels of lipid peroxides, thiobarbituric acid reactive substances, carbonyl proteins and the activities of some antioxidant enzymes were measured in brain, liver, kidney and skeletal muscle of goldfish, Carassius auratus L., over a time course of 3-12 h of hyperoxia exposure followed by 12 or 36 h of normoxic recovery. Exposure to high oxygen resulted in an accumulation of protein carbonyls in tissues throughout hyperoxia and recovery whereas lipid peroxides and thiobarbituric acid reactive substances accumulated transiently under short-term hyperoxia stress (3-6 h) but were then strongly reduced. This suggests that hyperoxia stimulated an enhancement of defenses against lipid peroxidation or mechanisms for enhancing the catabolism of peroxidation products. The activities of principal antioxidant enzymes, superoxide dismutase and catalase, were not altered under hyperoxia but catalase increased during normoxic recovery; activities may rise in anticipation of further hyperoxic excursions. In most tissues, the activities of glutathione-utilizing enzymes (glutathione peroxidase, glutathione-S-transferase, glutathione reductase) as well as glucose-6-phosphate dehydrogenase, were not affected under hyperoxia but increased sharply during normoxic recovery. Correlations between some enzyme activities and oxidative stress markers were found, for example, an inverse correlation was seen between levels of thiobarbituric acid reactive substances and glutathione-S-transferase activity in liver and catalase and glucose-6-phosphate dehydrogenase in kidney. The results suggest that liver glutathione-S-transferase plays an important role in detoxifying end products of lipid peroxidation accumulated under hyperoxia stress.     

Endogenous antioxidant defence system in rat liver following mercury chloride oral intoxication

Mercury is a highly toxic metal which induces oxidative stress. Superoxide dismutases, catalase, and glutathion peroxidase are proteins involved in the endogenous antioxidant defence system. In the present study rats were administered orally, by gavage, a single daily dose of HgCl2 for three consecutive days. In order to find a relation between the proteins involved in the antioxidant defence and mercury intoxication, parameters of liver injury, redox state of the cells, as well as intracellular protein levels and enzyme activities of Mn-dependent superoxide dismutase (MnSOD), Cu-Zn-dependent superoxide dismutase (CuZnSOD), catalase, and glutathione peroxidase (GPx) were assayed both in blood and in liver homogenates. HgCl2 at the doses of 0.1 mg/kg produced liver damage which that was detected by a slight increase in serum alanine aminotransferase and gamma glutamyl transferase. Hepatic GSH/GSSG ratio was assayed as a parameter of oxidative stress and a significant decrease was detected, as well as significant increases in enzyme activities and protein levels of hepatic antioxidant defence systems. Changes in both MnSOD and CuZnSOD were parallel to those of liver injury and oxidative stress, while the changes detected in catalase and GPx activities were progressively increased along with the mercury intoxication. Other enzyme activities related to the glutathione redox cycle, such as glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH), also increased progressively. We conclude that against low doses of mercury that produce a slight oxidative stress and liver injury, the response of the liver was to induce the synthesis and activity of the enzymes involved in the endogenous antioxidant system. The activities of all the enzymes assayed showed a rapidly induced coordinated response.     

Study on the antioxidant status of vitiligo patients of different age groups in Baroda

One of the major hypotheses in the pathogenesis of vitiligo is the oxidative stress hypothesis. Pollution plays a major role in the production of free radicals. Gujarat, a highly industrialized state in India has a high prevalence of vitiligo patients. No previous studies were done on the age-dependent antioxidant status of vitiligo patients in Baroda city, Gujarat. Blood samples were collected from vitiligo patients of different age groups (5-15, 16-25, 26-35, 36-45 yr) and from age matched healthy volunteers. Antioxidant enzymes in blood such as catalase, superoxide dismutase, glutathione peroxidase and non-enzymatic antioxidants such as reduced glutathione and plasma vitamin E were estimated. Lipid peroxidation levels in erythrocytes and the reducing equivalent system, i.e. glucose-6-phosphate dehydrogenase were also measured. Significant increase in superoxide dismutase activity and lipid peroxidation levels in erythrocytes was observed in all age groups of vitiligo patients as compared with age-matched healthy controls, wherein an increase of 55% (P < 0.02) was observed in superoxide dismutase activity and lipid peroxidation levels in 36-45 yr age group. Whole blood glutathione levels, erythrocyte glutathione peroxidase and glucose-6-phosphate dehydrogenase activity were decreased significantly, whereas erythrocyte catalase activity and plasma vitamin E levels were not different in vitiligo patients as compared with age-matched healthy controls. No specific age group showed a significant difference. This is the first report on the age-dependent antioxidant status of vitiligo patients in Baroda. The disease affects individuals of any age group as shown in this study and systemic oxidative stress might precipitate the pathogenesis of vitiligo in susceptible patients.     

Band 3 tyr-phosphorylation in normal and glucose-6-phospate dehydrogenase-deficient human erythrocytes

Haemolysis is usually episodic in glucose-6-phosphate dehydrogenase (G6PD) deficiency, often triggered by a period of oxidative stress. In the present work, we investigate a possible biochemical mechanism underlying the enhanced susceptibility of G6PD deficient red blood cells (RBC) to oxidative stress. We analysed eight male subjects with Mediterranean glucose-6P-dehydrogenase deficiency (G6PDd), class II, for their ability in phosphorylating erythrocyte membrane band 3 following oxidative and osmotic stress. Our findings show that this sensitivity is connected to an early membrane band 3 Tyr-phosphorylation in the presence of diamide. However, since both Syk, and Lyn kinases, and SHP-2 phosphatase, mostly implicated in the band 3 P-Tyr level regulation, are alike in content and activity in normal and patient erythrocytes, an alteration in the membrane organization is likely the cause of the anomalous response to the oxidant. We report, in fact, that hypertonic-induced morphological change in G6PDd erythrocyte induces a higher membrane band 3 Tyr-phosphorylation, suggesting a pre-existing membrane alteration, likely due to the chronic lowering of the redox systems in patients. We also report that 1-chloro-2,4-dinitrobenzene-pre-treatment of normal red cells can alter the normal protein-protein and protein-membrane interaction under hypertonic rather than oxidative stress, thus partially resembling the response in patients, and that RBC may utilize a wider range of redox defence, under oxidative conditions, including, but not exclusively, NADPH and glutathione. On the whole, these results would encourage a different approach to the evaluation of the effects of pharmacological administration to patients, giving more attention to the possible drug-induced membrane alteration evidenced by the abnormal band 3 Tyr-phosphorylation.     

Influence of very low doses of ionizing radiation on Synechococcus lividus metabolism during the initial growth phase

Previous results from this laboratory have shown that very low chronic doses of gamma radiation can stimulate proliferation of the Cyanobacterium Synechococcus lividus. This modification of cell proliferation occurred during the first doubling. In this paper, we have compared the metabolism of cells cultivated in a normal environment or under chronic irradiation. Incubation of the cells in a new medium induced a high superoxide dismutase (EC 1.15.1.1, SOD) activity at the 18th hour and a degradation of phycocyanin, thus demonstrating that cells were submitted to a photooxidative stress. This increase in superoxide dismutase activity was followed by concomittant peaks of glutathione reductase (EC 1.6.4.2, GR) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49, G6P-DH) at the 24th hour. Irradiated cultures at a dose of 53.5 mGray/year show an earlier and higher peak of SOD, GR, and G6P-DH. In a second stage, cultures showed an earlier onset of photosynthesis under irradiation, as evidenced by an increase in pigment content and an enhancement of glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13, GAP-DH). These results show that the radiostimulation is related to the activation of enzymes protecting against peroxides that were induced under oxidative circumstances and to the activation of a glucose catabolism via the oxidative pentose phosphate pathway.Abbreviations mGy milli-Gray - SOD superoxide dismutase - G6P-DH glucose-6-phosphate dehydrogenase - GAP-DH glycer-aldehyde-3-phosphate dehydrogenase - GSSG oxidized glutathione     

The NAD+ precursors, nicotinic acid and nicotinamide upregulate glyceraldehyde-3-phosphate dehydrogenase and glucose-6-phosphate dehydrogenase mRNA in Jurkat cells

To better understand the role of nicotinic acid and nicotinamide in the regulation of the oxidative stress response, we measured the levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and glucose-6-phosphate dehydrogenase (G6PD) mRNA in Jurkat cells treated with these NAD+ precursors. We used a modified nonradioactive Northern blot method and detected the mRNA using 18-mer digoxigenin (DIG)-labeled oligonucleotides as probes. We observed increased levels of the mRNAs for the two enzymes in treated cells. Our findings suggest that the NAD+ precursors may protect against oxidative stress and DNA damage by up-regulating the stress response genes GAPDH and G6PD.     

Age-related changes of antioxidant enzyme activities, glutathione status and lipid peroxidation in rat erythrocytes after heat stress

The aim of this study was to determine whether exposure to heat stress would lead to oxidative stress and whether this effect varied with different exposure periods. We kept 1-, 6- and 12-month-old male Wistar rats at an ambient temperature of either 22 degrees C or 40 degrees C for 3 and 7 days and measured glucose-6-phosphate dehydrogenase (G-6-PD), Cu,Zn-superoxide dismutase (Cu,Zn-SOD), catalase (CAT), selenium-dependent glutathione peroxidase (Se-GSH-Px) and glutathione-S-transferase (GST) activities and levels of thiobarbituric acid-reactive substances (TBARS), reduced glutathione (GSH) and oxidized glutathione (GSSG) in erythrocytes and determined GSH/GSSG ratio, total glutathione and the redox index. G-6-PD and CAT activities were found to be significantly increased in 1- and 6-month-old rats after 3 and 7 days of heat stress, but G-6-PD activities decreased in 12-month-old rats. Cu, Zn-SOD activity decreased in 1-month-old rats after heat stress, whereas it increased in 6- and 12-month-old rats. GST activity increased in all groups. GSH and total GSH levels and GSH/GSSG ratios decreased in 1- and 6-month-old rats but they increased in 12-month-old rats after heat stress. GSSG levels increased in 1- and 6-month-old rats but decreased in 12-month-old rats after heat stress. TBARS levels increased in all groups. Seven days of stress is more effective in altering enzyme activities and levels of GSH, GSSG and TBARS. When the effects of both heat stress and aging were examined together, it was interesting to note that they mostly influenced G-6-PD activity.