Energy production and redox status of rat red blood cells after reticulocytosis induced by various treatments.

Stimulated erythropoiesis and reticulocytosis can be induced by daily bleeding, or by phenylhydrazine (PHZ) treatment. We compared the in vivo effects of PHZ and bleeding treatment on haematological, energy and redox status parameters in red blood cells (RBC) of rats. The results showed that all followed haematological parameters were significantly lower in bleeding, compared to PHZ-treated rats. PHZ induced even 2.58-fold higher reticulocytosis as compared to bleeding treatment. Although PHZ induced higher reticulocytosis, respiration intensity and energy production was lower than in bleeding-induced reticulocytes. These alterations were the consequence of increased superoxide anion and peroxynitrite concentrations in PHZ-treated rats. Bleeding treatment resulted in increased activity of an antioxidative enzyme, superoxide dismutase. In conclusion, differences in these two experimental models for reticulocytosis may be used as tools for appropriate pharmacological testing of redox-active substances considering energy and redox processes, as well as apoptosis pathways.

Effect of chronic cadmium exposure on antioxidant defense system in some tissues of rats: protective effect of selenium.

The effects of selenium (Se) on antioxidant defense system in liver and kidneys of rats with cadmium (Cd)-induced toxicity were examined. Cd exposure (15 mg Cd/kg b.m./day as CdCl(2) for 4 weeks) resulted in increased lipid peroxidation (LP) in both organs (p<0.005 and p<0.01). Vitamin C (Vit C) was decreased in the liver (p<0.005), whereas vitamin E (Vit E) was increased in the liver and kidneys (p<0.005 and p<0.05) of Cd-exposed animals. Superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were decreased in both tissues (p<0.05 and p<0.005), whereas catalase (CAT) activity was decreased only in liver (p<0.005). Glutathione S-transferase (GST) increased in both tissues (p<0.005 and p<0.01). Treatment with Se (0.5 mg Se/kg b.m./day as Na(2)SeO(3) for 4 weeks) significantly increased liver and kidneys SOD and GSH-Px activities (p<0.05 to p<0.005), as well as CAT and GST activities only in the liver (p<0.01). In animals exposed to Se, both the concentrations of Vit C (p<0.01) and Vit E (p<0.005) were increased in both tissues. Co-treatment with Se resulted in reversal of oxidative stress with significant decline in analyzed tissues Cd burden. Our results show that Se may ameliorate Cd-induced oxidative stress by decreasing LP and altering antioxidant defense system in rat liver and kidneys and that Se demonstrates the protective effect from cadmium-induced oxidative damage.

The effects of nitroglycerine on the redox status of rat erythrocytes and reticulocytes.

The effects of nitroglycerine (NTG) are mediated by liberated nitric oxide (NO) after NTG enzymatic bio-transformation in cells. The aim of this study was to evaluate some products of NTG bio-transformation and their consequences on the redox status of rat erythrocytes and reticulocytes, considering the absence and presence of functional mitochondria in these cells, respectively. Rat erythrocyte and reticulocyte-rich red blood cell (RBC) suspensions were aerobically incubated (2 h, 37 degrees C) without (control) or in the presence of different concentrations of NTG (0.1, 0.25, 0.5, 1.0 and 1.5 mM). In rat erythrocytes, NTG did not elevate the concentrations of any reactive nitrogen species (RNS). However, NTG robustly increased concentration of methemoglobin (MetHb), suggesting that NTG bio-transformation was primarily connected with hemoglobin (Hb). NTG-induced MetHb formation was followed by the induction of lipid peroxidation. In rat reticulocytes, NTG caused an increase in the levels of nitrite, peroxinitrite, hydrogen peroxide, MetHb and lipid peroxide levels, but it decreased the level of the superoxide anion radical. Millimolar concentrations of NTG caused oxidative damage of both erythrocytes and reticulocytes. These data indicate that two pathways of NTG bio-transformation exist in reticulocytes: one generating RNS and the other connected with Hb (as in erythrocytes). In conclusion, NTG bio-transformation is different in erythrocytes and reticulocytes due to the presence of mitochondria in the latter.

The effect of coenzyme Q10 on blood ascorbic acid, vitamin E, an lipid peroxide in chronic cadmium intoxication.

The aim of our study was to investigate the possible protective role of coenzyme Q10 (CoQ10) administration on ascorbic acid (AsA), vitamin E (vit E), and lipid peroxide (LP) concentrations in the blood of rats chronically treated with cadmium. Results were compared to those obtained in control animals, as well as to those obtained in animals treated with olive oil. Compared to that of the control animals, the AsA concentration was significantly increased in rats treated with CoQ10 and olive oil, whereas vit E concentration was significantly increased in animals treated with cadmium, CoQ10, or cadmium + CoQ10. A significant decrease in LP concentration was noted in animals treated with cadmium or with cadmium + CoQ10o, whereas a significant increase was seen in animals treated with olive oil. Compared to that of the animals treated with olive oil, the ascorbic acid concentration was significantly decreased in rats treated with cadmium or with cadmium + CoQ10, whereas vit E concentration was significantly increased in animals treated with cadmium, CoQ10, or cadmium + CoQ10. LP concentration was significantly decreased in rats treated with cadmium, CoQ10, or cadmium + CoQ10. Our study showed that CoQ10 administration in rats chronically exposed to exogenous cadmium exerts beneficial effects on the nonenzymatic components of the antioxidant defense system, such as AsA and vit E, resulting in a decreased concentration of LP in the blood.

Effects of nitric oxide donors on energy metabolism of rat erythrocytes.

It is known that nitric oxide (NO) influences the activities of glycolytic enzymes, resulting in alteration of glycolysis rate. We investigated the effect of NO donors on the energy metabolism of rat erythrocytes. Rat erythrocyterich blood suspensions were aerobically incubated for 2 hours (1) as controls or (2) with different concentrations of NO donors: nitroglycerin (NTG), isosorbide dinitrate (ISDN), molsidomine (MO), and sodium nitroprusside (SNP). NTG, ISDN, MO, and SNP significantly (p < 0.05) increased glucose consumption and lactate accumulation in rat erythrocytes in a dose-dependent manner, indicating stimulation of glycolysis. The increased rate of glycolysis was accompanied by elevation of energy production (p < 0.05), but no changes in ATP levels were observed. The dose-dependent increase of glycolytic ATP production and the unaltered levels of ATP resulted in considerably shortened ATP-turnover time with the maximal experimental doses of NO donors used, indicating the stimulation of the ATP-consuming process in rat erythrocytes. The metabolic effects of NTG, ISDN, MO, and SNP were not mimicked by exogenous 8-Br-cGMP, NaNO2, or NaNO3, suggesting that the NO donor-induced stimulation of glycolysis and abbreviation of ATP-turnover time in rat erythrocytes was mediated by NO as an effector molecule, irrespective of the irreversible scavenger effect of hemoglobin. The implications of the NO effect on energy metabolism of erythrocytes is discussed.

The effect of cadmium and selenium on the antioxidant enzyme activities in rat heart.

Two month-old Wistar male albino rats were exposed during a 30-day period to a daily oral intake ad libitum of either 200 microg/mL Cd (as CdCl2), 0.1 microg/mL Se (as Na-selenite), or the same dosages of Cd + Se in drinking water. The daily intake from the water was calculated to be 15 mg Cd/kg and 7 microg Se/kg. Cadmium (Cd) accumulates in the heart (p < 0.005) and, in rats, decreases both body mass growth (p < 0.005) and heart mass (p < 0.02). Selenium (Se) significantly decreases the negative effect of Cd on body mass growth. In the hearts of Cd-treated rats, cadmium caused the decrease (p < 0.05) of selenium-dependent glutathione peroxidase (GSH-Px, EC 1.11.1.9) activity. At the same time, the activities of total superoxide dismutase (total SOD, EC 1.15.1.1), manganese-containing superoxide dismutase (Mn SOD), and copper-zinc-containing superoxide dismutase (CuZn SOD) were increased (p < 0.005). The activities of total SOD, CuZn SOD (p < 0.005), GSH-Px (p < 0.02), and glutathione-S-transferase (GST, p < 0.005) were increased in the hearts of Se-treated rats. However, by concomitant administration of Cd and Se, these changes were diminished (total SOD, GST) or were completely eliminated (Mn SOD, GSH-Px). These results indicate that Se only partly diminishes the effects of Cd cardiotoxicity.

The effects of selenium on the antioxidant defense system in the liver of rats exposed to cadmium.

Total superoxide dismutase (total SOD), copper zinc containing superoxide dismutase (CuZn SOD), and manganese superoxide dismutase (Mn SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and glutathione-S-transferase (GST) activities as well as ascorbic acid (AsA), and vitamin E (vit E) concentrations were analysed in the liver of rats exposed to cadmium (15 mg Cd/day/kg), selenium (7 micrograms Se/day/kg), and to cadmium+selenium (15 mg Cd + 7 micrograms Se/day/kg), and in control animals. Cadmium caused a decrease of total SOD, Mn SOD, CAT and GSH-Px but an increase of GST activity in the liver of rats. Contrary to cadmium, selenium caused a significant increase of the activity of these enzymes except for GSH-Px. By concomitant exposure to both cadmium and selenium, the toxic effects of cadmium on the activity of mentioned enzymes we abolished. In all exposed groups, the activity of enzyme glutathione-S-transferase was enhanced, indicating its increased role in prevention of lipid peroxidation. Cadmium decreased the concentration of AsA and increased the concentration of vitamin E in the liver, while selenium increased the concentration of both vitamins. However, by concomitant administration of cadmium and selenium, these changes were diminished and tended to reach control values.

Effects of nitric oxide donor, isosorbide dinitrate, on energy metabolism of rat reticulocytes.

Since nitric oxide (NO) in many cells is involved in energy metabolism, the aim of this study was to evaluate the role of isosorbide dinitrate (ISDN), a NO donor, in energy metabolism of rat reticulocytes, particularly due to their high content of hemoglobin--an effective scavenger of NO. Rat reticulocyte-rich red blood cell suspensions were aerobically incubated in the absence (control) or in the presence of different concentrations of ISDN. ISDN decreased total and coupled oxygen consumption (p<0.05) while increased uncoupled oxygen consumption (p<0.05) in a dose- and time-dependent manner. This was followed by enhancement of glycolysis, as measured by increased glucose consumption and lactate accumulation (p<0.05). Levels of all glycolytic intermediates in the presence of ISDN indicate only stimulation of pyruvate kinase activity. ISDN did not alter the concentration of ATP, while increased ADP and AMP levels (p>0.05). In rat reticulocytes under steady-state conditions, 95.4% of overall energy was produced by oxidative phosphorylation but only 4.6% by glycolysis. Due to a reduced coupled oxygen consumption in the presence of ISDN, ATP production via oxidative phosphorylation was significantly diminished. A simultaneous increase of glycolytic ATP production is not enough to ensure constant ATP production. The calculated mean ATP turnover time was prolonged by 199% in the presence of 1.5 mmol/l ISDN. In conclusion, ISDN a) inhibited total and coupled respiration but enhanced uncoupled respiration, b) stimulated glycolysis, c) decreased ATP production and d) prolonged ATP turnover time in rat reticulocytes. These effects were mediated by NO as the effector molecule.

Activities of superoxide dismutase and catalase in erythrocytes and plasma transaminases of goldfish (Carassius auratus gibelio Bloch.) exposed to cadmium.

Four groups of goldfish were exposed to cadmium in a concentration of 20 mg Cd/l water under aquarium conditions. The duration of exposure was 1, 4, 7 and 15 days. It was shown that the activity of superoxide dismutase (SOD) in the red blood cells (RBC) significantly decreased after the first day of cadmium exposure. However, the SOD activity increased after 7 and 15 days of cadmium treatment. Elevated activity of catalase (CAT) was found in erythrocytes of cadmium-treated fishes after 15 days, whereas plasma GOT levels was increased after 7 and 15 days and GPT levels after 1, 4, 7 and 15 days of cadmium treatment. This was accompanied by a significant decrease of blood hemoglobin concentrations (after 15 days) and hematocrit values (after 7 and 15 days). However, the concentration of blood glucose significantly increased after 1, 4, 7 and 15 days of cadmium exposure. These results indicate that cadmium causes oxidative stress and tissue damage in the exposed fishes.

Activities of superoxide dismutase and catalase in erythrocytes and transaminases in the plasma of carps (Cyprinus carpio L.) exposed to cadmium.

Total superoxide dismutase (SOD, EC 1.15.1.1) and catalase (CAT, EC 1.11.1.6) activities in erythrocytes and the glutamic acid-oxalacetic acid-transaminase (GOT, EC 2.6.1.1) and glutamic acid-pyruvic acid-transaminase (GPT, EC 2.6.1.2) activities in the plasma were measured in experimental groups of carps (Cyprinus carpio L.) exposed to cadmium in a concentration of 20 mg Cd/l water under aquarium conditions for 6, 12, 18 and 24 hours and in control fishes. It was shown that the total activity of SOD in the erythrocytes is significantly decreased after 12, 18 and 24 hours of cadmium exposure. Increased activities of CAT (after 24 hours) in the erythrocytes and GOT and GPT in the plasma were found in cadmium-treated fishes. At the same time the concentration of blood haemoglobin and haematocrit values were significantly diminished. These results indicate that cadmium causes oxidative stress and tissue damage in the exposed fishes.

The activities of superoxide dismutase, catalase and ascorbic acid content in the liver of goldfish (Carassius auratus gibelio Bloch.) exposed to cadmium.

The goldfish (Carassius auratus gibelio Bloch.) were exposed to cadmium in the concentration of 20 mg Cd/l water under aquarium conditions for 1, 4, 7 and 15 days. After exposure to cadmium, the activities of superoxide dismutase (SOD) and catalase (CAT) were significantly decreased. At the same time, the liver ascorbic acid (AsA) content was increased.

Protective influence of vitamin E on antioxidant defense system in the blood of rats treated with cadmium.

The effects of acute exposure to cadmium (Cd) on the blood antioxidant defense system, lipid peroxide concentration and hematological parameters, as well as the possible protective role of vitamin E were studied. Male Wistar albino rats (3 months old) were treated with cadmium (0.4 mg Cd/kg b.m., i.p., 24 h before the experiment) or with vitamin E + Cd (20 IU Vit E/kg b.m., i.m., 48 h + 0.4 mg Cd/kg b.m., i.p., 24 h before the experiment). The hematological parameters were assessed: red blood cell counts, hematocrit value and hemoglobin concentration were significantly decreased in the blood of Cd-treated rats. Intoxication with cadmium was also followed by significantly increased lipid peroxide concentrations. We also observed increased activity of antioxidant defense enzymes: copper zinc containing superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glutathione-S-transferase as well as concentrations of non-enzymatic components of antioxidant defense system: reduced glutathione, vitamin C and vitamin E. Pretreatment with vitamin E exhibited a protective role on the toxic effects of cadmium on the hematological values, lipid peroxide concentration as well as on enzymatic and non-enzymatic components of antioxidant defense system.

Effects of exogenous donor of nitric oxide-sodium nitroprusside on energy production of rat reticulocytes.

The effects of the sodium nitroprusside (SNP), a nitric oxide (NO) donor clinically used in the treatment of hypertensive emergencies on the energy production of rat reticulocytes were investigated. Rat reticulocyte-rich red blood cell suspensions were aerobically incubated without (control) or in the presence of different concentrations of SNP (0.1, 0.25, 0.5, 1.0 mM). SNP decreased total and coupled, but increased uncoupled oxygen consumption. This was accompanied by the stimulation of glycolysis, as measured by increased glucose consumption and lactate accumulation. Levels of all glycolytic intermediates indicate stimulation of hexokinase-phosphofructo kinase (HK-PFK), glyceraldehyde 3-phosphate dehydrogenase (GAPD) and pyruvate kinase (PK) activities in the presence of SNP. Due to the decrease of coupled oxygen consumption in the presence of SNP, ATP production via oxidative phosphorylation was significantly diminished. Simultaneous increase of glycolytic ATP production was not enough to provide constant ATP production. In addition, SNP significantly decreased ATP level, which was accompanied with increased ADP and AMP levels. However, the level of total adenine nucleotides was significantly lower, which was the consequence of increased catabolism of adenine nucleotides (increased hypoxanthine level). ATP/ADP ratio and adenylate energy charge level were significantly decreased. In conclusion, SNP induced inhibition of oxidative phosphorylation, stimulation of glycolysis, but depletion of total energy production in rat reticulocytes. These alterations were accompanied with instability of energy status.

Effect of cadmium and selenium on the antioxidant defense system in rat kidneys.

To examine effects of exogenous Cd on the kidney antioxidant defense system (AOS) and the possible protective role of Se against Cd toxicity, male Wistar albino rats (2 months old) were exposed during 30 days to oral intake of 200 ppm Cd (as CdCl2), 0.l ppm Se (as Na-selenite) or to the same doses of Cd / Se, simultaneously. Marked accumulation of Cd (23.44 +/- 0.69 micrograms/g w.m.) and marked alterations of AOS, resulting in kidney injury (renal pseudohypertrophy), were found in Cd-treated rats. Activities of total superoxide dismutase (SOC, EC 1.15.1.1), manganese-containing superoxide dismutase (MnSOD) and selenium-dependent glutathione peroxidase (Se GSH-Px, EG 1.11.1.9) were significantly reduced, whereas that of glutathione-S-transferase (CST, EC 2.5.1.18) and vitamin E (vit E) concentration were significantly increased in the kidneys of Cd-treated rats. Kidney catalase (CAT, EC 1.11.1.6) activity, ascorbic acid (AsA) and red blood cell glutathione (GSH, GSSG) levels were not markedly influenced by CD uptake. In kidneys of Se treated rats, the activities of total SOD, copper-zinc-containing superoxide dismutase (CuZnSOD) and GST were significantly increased Activities of kidney CAT and Se GSH-Px were largely unchanged, whereas significant increases of the kidney AsA and vit E concentrations occurred. In Cd + Se-cotreated rats, the kidney activities of MnSOD, CAT and Se GSH-Px, as well as vit E concentration, were the same as in controls, whereas CuZnSOD and GST activities and concentration of AsA exceeded normal values. These data indicate that Se only partially improves the AOS that is insufficient to prevent Cd-induced nephrotoxicity.

Cadmium-induced changes of antioxidant and metabolic status in red blood cells of rats: in vivo effects.

Chronic exposure of adult rats to dietary intake of cadmium (15 mg CdCl2/day/kg for 30 days) leads to development of anemia and thrombocytosis. Anemia is characterized by significant reticulocytosis (13.1 +/- 1.0%), anysocytosis, poikilocytosis, iron deficiency and marked alterations of antioxidant and metabolic status of red blood cells. Activities of SOD, catalase, GPx and GR were significantly increased in red blood cells of cadmium-treated rats. In treated animals cadmium induced an increase of red cell reduced and oxidized glutathione with no changes of GSSG/GSH ratio. However, significant reduction of lipid peroxidation was found. Plasma levels of tocopherol and ascorbate, as well as activity of glutathione-S-transferase, were all significantly increased in cadmium-treated rats. The energy metabolism of red blood cells was deeply altered in cadmium-treated rats. The levels of ATP, ADP, AMP and TAN were significantly increased while ATP/ADP ratio and adenylate energy charge (AEC) were significantly reduced. The level of 2,3-BPG was somewhat lower, but 2,3-BPG/Hb ratio was considerably higher, in red blood cells of cadmium-treated rats.