Influence of indole acetic acid on antioxidant levels and enzyme activities of glucose metabolism in rat liver

Indole acetic acid (IAA) is an auxin and can be synthesized in animals. This compound is metabolized in vitro by peroxidase, producing reactive oxygen species. The toxic effect of indole acetic acid in leukocytes is associated with peroxidase activities and these processes have been implicated in activation of glucose and glutamine metabolism. However, studies in vitro have shown that IAA, in absence of peroxidase, is an antioxidant almost as high in potency as those of other indolic compounds. The purpose of this study was to investigate the possible involvement of a toxic effect of indole acetic acid in the liver, as evidenced by oxidative stress and enzyme activities of the glucose pathway. The animals received IAA by subcutaneous or gavage administration in a phosphate buffered saline (the control group received only the phosphate buffered saline). The other groups received IAA at concentrations of 1 mg, 18 mg and 40 mg per kg of body mass per day. Treatments with 18 mg and 40 mg IAA decreased the activity of catalase by both subcutaneous (30% and 26%) or gavage administration (19% and 28%), respectively. A similar effect was observed on the activity of glutathione peroxidase of animals exposed to 18 mg and 40 mg IAA: A decrease of 34% and 29%, respectively, for subcutaneous administration and a decrease of 29% and 25%, respectively, for gavage administration. However, in neither source of administration did the acid alter superoxide dismutase, glutathione reductase and myeloperoxidase activities. Another alteration was observed in respect of reduced glutathione content in this organ. The lipid peroxidation level showed a significant decrease with subcutaneous (30%, 29% and 24%) and gavage administration (25%, 26% and 24%) using 1 mg, 18 mg and 40 mg of IAA, respectively compared with the control. The reduced glutathione content and catalase activity in the plasma were not altered by either of the two methods of administration. In addition to these findings, after subcutaneous or gavage administration of IAA, the activities of hepatic enzymes of glucose metabolism were not affected (glucokinase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase and citrate synthase). Evidence is presented herein that IAA did not have a pro-oxidant effect in the liver as deduced from a reduction of catalase and glutathione peroxidase activities, a decrease of lipid peroxidation content and no alteration of the pool of reduced glutathione. The effects of IAA were independent of the way of administration.     

Cyclosporin A generates superoxide in smooth muscle cells

Cyclosporin A (CsA) generates superoxide in smooth muscle cells. Our earlier studies have demonstrated that the increase in the vasopressin type 1 receptor induced in vascular smooth muscle cells in the presence of CsA is probably due to superoxide (Krauskopf et al., J Biol Chem 278, 41685-41690, 2003). This increase in vasopressin receptor is likely at the base of increased vascular responsiveness to vasoconstrictor hormones and hypertension induced by CsA. Here, we demonstrate that CsA produces superoxide. In addition, our data show that superoxide generation does not originate from the major cellular superoxide generating systems NAD(P)H oxidase or xanthine oxidase. Our results suggest that the side effects of CsA could be diminished with the help of SOD mimetic drugs.     

Extracellular ATP through P2 receptors activates AMP-activated protein kinase and suppresses superoxide generation in cultured mouse podocytes

Podocytes are an important constituent of the glomerular filtration barrier. The function of these glomerular cells is affected by extracellular nucleotides through P2 receptors. The activation of P2 receptors may lead to the activation of NAD(P)H oxidase, the key enzyme in oxidative stress, with the intracellular pathways leading to intracellular ATP depletion associated with an increase in the intracellular AMP:ATP ratio. This deregulation of the energy balance activates AMP-activated protein kinase (AMPK) to restore energy homeostasis. We investigated whether P2 receptor activation influences NAD(P)H oxidase-dependent rate of superoxide anion (O₂^•−) generation and AMPK activity in cultured mouse podocytes. The rate of O₂^•− generation was measured by chemiluminescence and changes in AMPK activity were determined by immunoblotting against AMPKα-Thr¹⁷²-P. The addition of 100 μM ATP induced a rapid and transient decrease in rate of O₂^•− generation and increased AMPK phosphorylation with maximal effects in the first minute (2.44 ± 0.09 versus 1.62 ± 0.06 nmol/mg protein/min, P < 0.05 and 0.64 ± 0.04 versus 0.97 ± 0.07, P < 0.05, respectively). Both parameters returned to control levels at 10 min. Suramin (300 μM, P2 receptor antagonist) and compound C (100 μM, AMPK inhibitor) completely, and STO-609 (25 μM, CaMKK-β inhibitor) partially, prevented ATP action in rate of O₂^•− generation and AMPK phosphorylation. Various ATP analogues (10 μM) mimicked the effects of ATP on rate of O₂^•− generation and AMPK phosphorylation. The data indicate that extracellular ATP, acting through P2 receptors upstream of CaMKK-β, modulates podocyte function through simultaneous effects on AMPK and NAD(P)H oxidase activities. This mechanism may play a role in restoring energy homeostasis after oxidative stress.     

Nox-generated ROS modulate glucose uptake in a leukaemic cell line

The discovery of superoxide-generating enzymes homologues of phagocytic NAD(P)H oxidase, the Nox family, has led to the concept that reactive oxygen species (ROS) are ‘intentionally’ generated with biological functions in various cell types. In this study, by treating an acute leukaemic cell line with different antioxidants, ROS generation was shown to be crucially involved in the modulation of glucose transport (mediated by Glut1), which is frequently up-regulated in cancer cells. Then, this study tried to elucidate ROS source(s) and mechanisms by which ROS are involved in Glut1 activity regulation. Results prove that Nox2 and Nox4 are the candidates and that phosphorylation processes are important in the regulation of glucose uptake on which cancer cells rely. On the whole, data suggest that both Glut1 and Nox homologues may be considered new potential targets in the treatment of leukaemia.     

Percentage of phagocytosis,production of O2·−, H2O2 and NO,and antioxidant enzyme activities of rat neutrophils in culture

Changes in the integrity, ultrastructure, phagocytosis capacity, and production of H₂O₂, O₂^{· −}and NO₂⁻ were evaluated in cultured neutrophils. The activities of the antioxidant enzymes (catalase—CAT, superoxide dismutase—SOD and glutathione-dependent peroxidase—GSH-Px) were measured under similar conditions. The integrity of the cells remained unchanged up to 18 h. After 24 h, the number of viable cells in culture dropped by 16 per cent. The percentage of viable cells in culture was of 72 per cent even after 72 h. An ultrastructural analysis of the cells was carried out after 3, 6, 12, 24, 48, and 72 h in culture. Neutrophils started developing morphologic changes after 24 h: decreased cell volume, abundant vacuoles (mainly around the nucleus), and also the presence of autophagic vacuoles. This period was then chosen for the study of neutrophil function and antioxidant enzyme activities. Neutrophils cultured for 24 h presented reduced phagocytosis capacity. The rates of production of H₂O₂ and O₂^{· −} remained unchanged after 24 h in culture. Concomitantly, these cells were also able to produce NO in significant amounts. The production of O₂^·− in response to PMA stimulus was lowered in 24-h cultured cells. Possibly, the production of oxygen and nitrogen reactive species accomplished with a decrease in the activities of CAT and GSH-Px play a key role for the process of apoptosis which takes place in neutrophils under these conditions. © 1998 John Wiley & Sons, Ltd.     

Effects of post-injury hypothermia and nerve growth factor infusion on antioxidant enzyme activity in the rat: implications for clinical therapies

The pathological sequelae of traumatic brain injury (TBI) include increased oxidative stress due to the production of reactive oxygen species (ROS). Regulation of ROS levels following TBI is determined primarily by antioxidant enzyme activity that in turn can be influenced by nerve growth factor (NGF). Hypothermia is one of the current therapies designed to combat the deleterious effects of TBI. However, it has been shown to suppress post-trauma increases in NGF levels in rat brain. The present study sought to determine whether post-injury hypothermia also impairs the antioxidant response to injury, and if such an effect could be reversed by infusion of exogenous NGF. We employed a lateral controlled cortical impact injury model in rat, followed by moderate hypothermia treatment with supplemental intracerebroventricular infusion of NGF or vehicle. The time course of changes in post-injury/intervention levels of NGF and activity of three major enzymes responsible for ROS scavenging, catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD), was determined in the hippocampus. Relative to levels in injured, normothermic animals, hypothermia treatment not only suppressed NGF levels, but also attenuated CAT and GPx activity, and increased SOD activity. Infusion of NGF in injured, hypothermia-treated animals was ineffective in restoring hippocampal antioxidant enzymes activity to levels produced after injury under normothermic conditions, although it was able to increase septal cholinergic (choline acetyltransferase) enzyme activity. These results have implications for clinical treatment of TBI, demonstrating that moderate hypothermia suppresses NGF and the antioxidant response after TBI; the latter cannot be countered by exogenous NGF administration.     

Oxidative stress responses during cassava post-harvest physiological deterioration

A major constraint to the development of cassava (Manihot esculenta Crantz) as a crop to both farmers and processors is its starchy storage roots' rapid post-harvest deterioration, which can render it unpalatable and unmarketable within 24–72 h. An oxidative burst occurs within 15 min of the root being injured, that is followed by the altered regulation of genes, notably for catalase and peroxidase, related to the modulation of reactive oxygen species, and the accumulation of secondary metabolites, some of which show antioxidant properties. The interactions between these enzymes and compounds, in particular peroxidase and the coumarin, scopoletin, are largely confined to the vascular tissues where the visible symptoms of deterioration are observed. These, together with other data, are used to develop a tentative model of some of the principal events involved in the deterioration process. Abbreviations: ACMV, African cassava mosaic virus; AFLP, amplified fragment length polymorphism; CAT, catalase; cDNA, complementary deoxyribonucleic acid; CIAT, International Centre for Tropical Agriculture; Cu/ZnSOD, copper/zinc superoxide dismutase; DAB, 3,3-diaminobenzidine tetrahydrochloride; DPPH, 1,1-diphenyl-2-picrylhydrazyl; FeSOD, iron superoxide dismutase; FW, fresh weight; GUS, -glucuronidase; HPTLC, high-performance thin-layer chromatography; HR, hypersensitive response; IEF-PAGE, isoelectric focusing polyacrylamide gel electrophoresis; MAS, marker-assisted selection; MeJa, methyl jasmonate; MnSOD, manganese superoxide dismutase; NADPH, nicotinamide adenine dinucleotide phosphate (reduced form); NBT, nitroblue tetrazolium; PAL, phenylalanine ammonia-lyase; PCD, programmed cell death; PCR, polymerase chain reaction; POX, peroxidase; PPD, post-harvest physiological deterioration; QTL, quantitative trait loci; ROS, reactive oxygen species; RT, room temperature; SAR, systemic acquired resistance; SDS, sodium dodecyl sulfate; SOD, superoxide dismutase     

Oxidative stress responses during cassava post-harvest physiological deterioration

A major constraint to the development of cassava (Manihot esculenta Crantz) as a crop to both farmers and processors is its starchy storage roots’ rapid post-harvest deterioration, which can render it unpalatable and unmarketable within 24–72 h. An oxidative burst occurs within 15 min of the root being injured, that is followed by the altered regulation of genes, notably for catalase and peroxidase, related to the modulation of reactive oxygen species, and the accumulation of secondary metabolites, some of which show antioxidant properties. The interactions between these enzymes and compounds, in particular peroxidase and the coumarin, scopoletin, are largely confined to the vascular tissues where the visible symptoms of deterioration are observed. These, together with other data, are used to develop a tentative model of some of the principal events involved in the deterioration process.     

Superoxide production and oxygen consumption in endothelium-intact and -denuded artery stimulated by angiotensin II

Arteries stimulated by angiotensin II (AII) to contract do not display the expected augmentation of O₂ consumption seen with other cardiovascular contractile agonists. We tested the hypothesis that superoxide (O₂⁻) or other reactive oxidant species generated by AII played a role in the paradoxical O₂ consumption response in porcine carotid artery, with or without an intact endothelium. Endothelium-denuded arteries were incubated with either 1 μM diphenylene iodonium (DPI), an inhibitor of NAD(P)H oxidase, 300 u/ml superoxide dismutase (SOD), a scavenger of O₂⁻, or 20 U/ml catalase, an enzyme which promotes conversion of O₂⁻ (scavenged in the form of H₂O₂) to O₂. DPI treatment resulted in the expected increase in O₂ consumption upon contractile activation with AII challenge (1.05± 0.23 μmol/g/min; n = 6, p < .01), as did treatment with SOD (0.67± 0.20 μmol/g/min; n = 4, p < .05). Catalase incubation resulted in a burst of O₂ generation upon AII challenge (1.30 ± 0.21 μmol/g/min; n = 10, p < .001). In endothelium-intact arteries, O₂ consumption was again not augmented with AII challenge; instead, a burst of O₂ production was observed (0.66 ± 0.22 μmol/g/min; n = 9, p < .05), which was not affected further by addition of catalase. Thus, the absence of apparent augmentation of O₂ consumption during contractile activation of endothelium-denuded arteries was attributed to simultaneous NAD(P)H oxidase-dependent production of O₂⁻, and attendant H₂O₂ and O₂ generation which either and masked the detection of O₂ consumed or suppressed mitochondrial uptake of O₂, or both. An intact endothelium was required to manifest the burst of O₂ generation with AII stimulation under normal conditions. (Mol Cell Biochem xxx: 235–239, 2005)     

NAD(P)H oxidase isoform Nox2 plays a prosurvival role in human leukaemia cells

The mechanism involved in the prosurvival effect of interleukin-3 on the human acute myeloid leukaemia cell line M07e is investigated. A decrease in intracellular reactive oxygen species (ROS) content, glucose transport activity and cell survival was observed in the presence of inhibitors of plasma membrane ROS sources, such as diphenylene iodonium and apocynin, and by small interference RNA for Nox2. Moreover, IL-3 incubation stimulated the synthesis of Nox2 cytosolic sub-unit p47phox and glucose transporter Glut1. Thus, the inhibition of ROS generation by Nox inhibitors stimulated apoptosis showing that ROS production, induced by IL-3 via Nox2, protects leukaemic cells from cell death. Also incubation with receptor tyrosine kinase inhibitors, such as anti-leukaemic drugs blocking the stem cell factor receptor (c-kit), showed similar effects, hinting that IL-3 transmodulates c-kit phosphorylation. These mechanisms may play an important role in acute myeloid leukaemia treatment, representing a novel therapeutic target.     

Nitric oxide treatment alleviates drought stress in wheat seedlings

The effects of sodium nitroprusside (SNP; nitric oxide donor) treatment on drought stress induced by PEG for different periods of time in wheat seedlings were investigated. Our results suggested that treatment for 2, 4 and 6 d with 15 % PEG could be termed as mild, moderate and severe stress, respectively. Drought stress induced accumulation of hydrogen peroxide and resulted in lipid peroxidation. On the other hand, activities of SOD, CAT and PAL increased under mild stress to counteract the oxidative injury and then decreased when the stress became severe (6 d). As the effect of SNP treatment, 0.2 mM enhanced wheat seedlings growth and kept high relative water content and alleviated the oxidative damage. However, 2 mM SNP aggravated the stress as a result of uncontrolled generation of reactive oxygen species and ineffectiveness of antioxidant systems.     

Studies on hepatic oxidative stress and antioxidant defence systems during arteether treatment of Plasmodium yoelii nigeriensis infected mice

Reactive Oxygen species play an important role in pathology during malaria infection. The status of hepatic oxidative stress and antioxidant defence indices was studied during Plasmodium yoelii nigeriensis (P. y. nigeriensis) infection in mice and arteether treatment of P. y. nigeriensis infected mice. P. y. nigeriensis infection caused a significant increase in hepatic xanthine oxidase, rate of lipid peroxidation, reduced glutathione (GSH) and glutathione reductase with progressive rise in parasitemia. This was accompanied by a significant decrease in hepatic superoxide dismutase (SOD) and catalase with increase in parasitemia. Arteether treatment (10 mg/kg body weight of mice) of infected mice from day 2 of post infection resulted in complete clearance of parasitemia on day 4 of post infection which was accompanied by restoration of all the oxidative stress and antioxidant defence indices to normal levels.     

Retinol supplementation induces oxidative stress and modulates antioxidant enzyme activities in rat Sertoli cells

Recent intervention studies revealed that supplementation with retinoids resulted in a higher incidence of lung cancer. Recently the causal mechanism has begun to be clarified. We report here that retinol caused cellular oxidative stress and modulated superoxide dismutase, catalase and glutathione peroxidase activities. Retinol (7 μM) significantly increased TBARS, conjugated dienes, and hydroperoxide-initiated chemiluminescence in cultured Sertoli cells. In response to retinol treatment superoxide dismutase, catalase and glutathione peroxidase activities increased. TBARS content and catalase activities were decreased by a free radical scavenger. These findings suggest that retinol may induce oxidative stress and modulate antioxidant enzyme activities in Sertoli cells.     

Bone marrow derived cells and reactive oxygen species in hypertrophy of contralateral kidney of transient unilateral renal ischemia-induced mouse

Abstract

Renal mass reduction, such as unilateral nephrectomy induces a compensatory hypertrophy of remaining renal mass in response to overload induced by reduction of functional renal parenchyma. In our recent study, we observed that the recovery of ischemic injured kidney following transient unilateral renal ischemia took longer time than that following transient bilateral renal ischemia, indicating that non-damaged kidney may affect the damaged kidney and vice versa. Here, we investigated whether transient and partial renal parenchymal injury by transient unilateral renal ischemia (UI) results in the hypertrophy of its contralateral kidney (CLK) and reactive oxygen species is associated with the hypertrophy. Thirty minutes of UI resulted in gradual increase in CLK weight over time. UI increased superoxide formation, but not lipid peroxidation in the CLK. After UI, a significant increase in the number of NADPH oxidase 2 (Nox2)-expressing cells and the level of Nox2 expression in the CLK was observed. In parallel with the increases in Nox2-expressing cells in CLKs, infiltration of bone marrow-derived cells (BMDC) increased in CLK. Treatments with Mn(III) Tetrakis(1-methyl-4-pyridyl) porphyrin (MnTMPyP, a superoxide dismutase (SOD) mimetic) and apocynin (a putative NADPH oxidase inhibitor) inhibited UI-induced hypertrophy of CLK along with reduction in Nox2-positive cell, BMDC, amount of Nox2 expression and superoxide formation. In conclusion, transient and partial renal mass reduction by UI resulted in the hypertrophy of CLK through increased ROS formation by infiltrated cells into the interstitium of CLK.     

H2O2 but not $${\hbox{O}_{2}^{-}}$$ elevated by oxidized LDL enhances human aortic smooth muscle cell proliferation

The oxidation of low density lipoprotein (LDL) as a key event in atherosclerosis suggests that antioxidant interventions may reduce the risk of atherosclerosis. However, the better strategies among antioxidant remedies for atherosclerosis remains difficult to be determined. Here, we show that oxidized LDL increases the steady-state level of intracellular hydrogen peroxide through stimulating the protein expressions of Nox1 and Cu/Zn superoxide dismutase (SOD) in human aortic smooth muscle cells (SMCs). The intracellular content of hydrogen peroxide rather than superoxide is a key modulator for vascular SMC (VSMC) proliferation, implying that without co-expression of catalase, increased Cu/Zn-SOD activity alone may not be beneficial to reduce the growth of VSMC in an atherosclerotic plaque.     

Protective effect of ozone treatment on the injury associated with hepatic ischemia-reperfusion: Antioxidant-prooxidant balance

The effects of ozone treatment on the injury associated to hepatic ischemia-reperfusion (I/R) was evaluated. Ozone treatment (1 mg/kg daily during 10 days by rectal insufflation) is shown to be protective as it attenuated the increases in transaminases (AST, ALT) and lactate levels observed after I/R. I/R leads to a decrease in endogenous antioxidant (SOD and glutathione) and an increase in reactive oxygen species (H₂O₂) with respect to the control group. However, ozone treatment results in a preservation (glutathione) or increase (SOD) in antioxidant defense and maintains H₂O₂ at levels comparable to those in the control group. The present study reports a protective effect of ozone treatment on the injury associated to hepatic I/R. The effectiveness of ozone could be related to its action on endogenous antioxidants and prooxidants balance in favour of antioxidants, thus attenuating oxidative stress.