Effects of glycerol-induced acute renal failure on tissue glutathione and glutathione-dependent enzymes in the rat

The activities of tissue glutathione (reduced and oxidized) and glutathione-dependent enzymes such as glutathione S-transferase (GSH S-transferase), glutathione reductase (GSSG reductase) and glutathione peroxidase (GSH-Px) were determined for control and uremic rats. Acute renal failure (ARF) was produced by glycerol-water injection. Cytosolic and microsomal GSH S-transferase activity in the kidney was decreased by 38% and 15%, respectively. Hepatic microsomal GSH S-transferase was also decreased by 40% in uremic rats. GSH-Px activity was decreased by 51% in the cytosolic fraction and 33% in the microsomal fraction in the kidney, but was not affected in the liver and whole blood. GSSG reductase activity was also decreased by 48% in the cytosolic fraction in the kidney of uremic rats. In whole blood, however, GSSG reductase activity was increased by 12-fold (0.66 +/- 0.12 mumol NADPH oxidized/min/ml blood in the control; 8.03 +/- 3.29 mumol NADPH oxidized/min/ml blood in uremia). Although the total glutathione concentrations were not significantly affected, the GSSG/GSH ratio, which is an indication of oxidative stress, was significantly increased in the liver and whole blood of uremic rats. In addition to the decreases in hepatic and renal GSH S-transferase activities, which is important in drug disposition, ARF caused decreases in GSSG reductase and GSH-Px activity, which are essential for the protection against lipid peroxidation.     

Kinetics and mechanism of the oxidation of the glutathione dimer by hypochlorous Acid and catalytic reduction of the chloroamine product by glutathione reductase

Oxidized glutathione (GSSG) reacts with two molar equivalents of HOCl/OCl- (a neutrophil-derived oxidant and a common biocide) to form the dichloro (bis-N-chloro-gamma-l-glutamyl) derivative (NDG). The reaction of less than two molar equivalents of HOCl with GSSG does not yield the unsymmetrical monochloro derivative (NCG) but rather a stoichiometric amount of NDG and GSSG. This result is explained by a faster reaction of the second equivalent of HOCl with NCG than that of the first equivalent of HOCl with GSSG. The rates of reaction of GSSG2-, GSSG3-, and GSSG4- (successive deprotonation of the ammonium groups) have been investigated, and it is clear that GSSG2- is unreactive, whereas GSSG4- is about twice as reactive as GSSG3-. Accordingly, the following mechanism is proposed (constants for 5 degrees C): H+ + OCl- = HOCl, pK1 = -7.47; GSSG2- = GSSG3- + H+, pK2 = 8.5; GSSG3- = GSSG4- + H+, pK3 = 9.5; GSSG3- + HOCl --> NCG3- + H2O, k4 = 2.7(2) x 106 M-1 s-1; GSSG4- + HOCl --> NCG4- + H2O, k5 = 3.5(3) x 107 M-1 s-1; NCG3- --> NDG4- + H+, k6 = fast; and NCG4- + HOCl --> NDG4- + H2O, k7 = fast. At physiologic pH, the k4 pathway dominates. NDG decomposes at pH 7.4 in a first-order process with kdec = 4.22(1) x 10-4 s-1 (t1/2 = 27 min). Glutathione reductase (EC 1.6.4.2) is capable of catalyzing the reduction of NDG by NADPH. The only NDG-derived product that is observed (by NMR) after the reduction by NADPH is GSH. Thus, in the presence of the GOR/NADPH system, GSH is capable of redox buffering a 3/2 mol equiv of HOCl rather than a 1/2 mol equiv as previously assumed.     

Changes in antioxidant defense systems induced by thiram in V79 Chinese hamster fibroblasts.

The role of antioxidant defence systems in protection against oxidative damage of lipids and proteins induced by fungicide thiram during in vitro exposure was investigated in cultured Chinese hamster V79 cells with normal, depleted, and elevated glutathione (GSH) levels. We analyzed the catalytic activities of superoxide dismutases (SOD1 and SOD2), Se-dependent and Se-independent glutathione peroxidases (GSH-Px), glutathione reductase (GR), and catalase (CAT), as well as total glutathione/glutathione disulfide ratio (GSH(total)/GSSG). Thiram treatment resulted in an increase in activities of SOD1, Se-dependent GSH-Px, and GR at the highest tested dose (150 microM). On the contrary, inhibition of CAT and Se-independent GSH-Px activities, and no significant changes in the level of SOD2 activity was observed at any tested doses (100-150 microM). GSH(total)/GSSG ratio in the 100 microM thiram treated cells was not significantly changed comparing to the control, despite significant decrease of GSH total (50%). In 150 microM thiram treated cells the ratio falls to 43% of control value. Pretreatment with l-buthionine sulfoximine (L-BSO), an inhibitor of GSH synthesis, significantly enhanced decrease in CAT and Se-independent GSH-Px activities, as well as GSH(total)/GSSG ratio, and reduced Se-dependent GSH-Px activity, following exposure to thiram. Simultaneously, L-BSO pretreatment enhanced increase in SOD1 activity, and had no effect on SOD2, following thiram exposure. Pretreatment with N-acetyl cysteine (NAC), a GSH precursor, prevented enzymatic changes in CAT, Se-dependent GSH-Px, GR, SOD1 activities, and significantly decreased SOD2 activity following exposure to thiram. GSH(total)/GSSG ratio was restored to the control value. This study suggests that following the changes in antioxidant defense systems thiram can act through the production of free radicals.     

A novel biologically active seleno-organic compound--III. Effects of PZ 51 (Ebselen) on glutathione peroxidase and secretory activities of mouse macrophages

PZ 51 (2-phenyl-1,2-benzisoselenazol-3(2H)-on), a selenium-containing compound with glutathione peroxidase (GSH-Px)-like activity, was administered to selenium-deficient mice for 5 days. A significant increase in peritoneal macrophage GSH-Px activity after treatment was only observed when basal GSH-Px activity was almost zero (i.e. in 19 weeks selenium-deficient animals), possibly due to binding of PZ 51 to the macrophages. This indicates that PZ 51 releases very little, if any, free selenium for incorporation into endogenous GSH-Px. The compound in vitro exerted a concentration-dependent inhibition of the generation of chemiluminescence by resident mouse peritoneal macrophages and a partial inhibition of the production of prostaglandin E2 by resident peritoneal macrophages. beta-Glucuronidase production by C. parvum-activated peritoneal macrophages was unaffected by PZ 51. These in vitro data can be explained on the basis of a selective peroxide scavenging and/or GSH-Px-like activity of PZ 51, offering a novel approach to anti-inflammatory therapy.     

The effects of bucillamine on glutathione and glutathione-related enzymes in the mouse

The effect of bucillamine (BA) on glutathione (GSH) and GSH-related enzymes was investigated in C57 mouse. Administration of high doses of BA (150-400 mg/kg) produced a dose-dependent depletion (20-44%) of hepatic GSH, which was similar in magnitude to that produced by equimolar doses of other sulphydryl drugs studied previously. GSH depletion after acute BA administration correlated well with the elevation of serum glutamic-pyruvic transaminase (SGPT) (6-9-fold increase above control). The increase in SGPT after chronic administration (7 days), although significantly higher than the controls, was however much less than after acute administration. The hepatic GSH concentrations of mice given 7 days of BA were similar to the controls, again correlating well with SGPT activity. Administration of BA (150-400 mg/kg) caused also a significant dose-dependent increase in the oxidized glutathione (GSSG) in blood by 2-7-fold, as well as a dose-dependent increase in blood glutathione S-transferase (GST) activity (2-13-fold). In an in vitro experiment, hepatic GST activity was activated by various concentrations of BA (1 microM-1mM). There was little or no effect on GSSG reductase and on glutathione peroxidase (GSH-Px) after acute administration of BA. Chronic administration of BA had no effect on hepatic GSSG reductase and GSH-Px, but GSSG reductase activity in blood was increased significantly by 4-fold. It is possible that BA may affect the redox status through auto-oxidation and oxidation with endogenous thiols such as glutathione, affecting GSH concentrations and the GSH/GSSG ratio in tissues and, thus, having both metabolic and toxicological consequences. Whether or not the induction of GST activity in vivo in blood and in vitro in liver enzyme preparations shared the same underlying mechanism(s) requires further investigation.     

Binding of two anthranilic acid derivatives to human albumin, erythrocytes, and lipoproteins: evidence for glafenic acid high affinity binding

The binding of two anthranilic acid derivatives, glafenic and floctafenic acids, to human erythrocytes and plasma proteins has been investigated in vitro by equilibrium dialysis. Despite their close chemical structures it was shown that the binding of the two compounds to serum albumin, lipoproteins, and erythrocytes was dramatically different both in quality and quantity. Using various techniques including fluorometry and circular dichroism, it was shown that glafenic acid binds to the human serum albumin (HSA) warfarin/azapropazone site and that floctafenic acid binds to both warfarin/azapropazone and benzodiazepine sites. Glafenic acid is strongly bound to HSA with n = 1, k = 2.4 X 10(6) liters/mol and to erythrocytes with N = 12.4 mumol/liter, K = 1.7 X 10(6) liters/mol. Floctafenic acid is bound with a weaker affinity to HSA, n = 2, k = 0.3 X 10(6) liters/mol and to erythrocytes, N = 2900 mumol/liter and K = 0.007 X 10(6) liters/mol.     

Ginsenoside-Rd attenuates oxidative damage related to aging in senescence-accelerated mice

Among the various theories of the aging process, the free radical theory, which proposes that deleterious actions of free radicals are responsible for the functional deterioration associated with aging, has received widespread attention. The theory suggests that enhancement of the antioxidative defence system to attenuate free-radical-induced damage will counteract the aging process. We used senescence-accelerated mice (SAM) to investigate the relationship between aging and the antioxidative defence system and evaluated the effects of ginsenoside-Rd, the saponin from ginseng, by measuring antioxidative defence system parameters, including the glutathione (GSH)/glutathione disulfide (GSSG) redox status, antioxidative enzyme activity and level of lipid peroxidation. SAM at 11 months of age (old SAM) showed a significantly lower hepatic GSH/GSSG ratio, due to decreased GSH and increased GSSG levels, than SAM at 5 weeks of age (young SAM). However, the administration of ginsenoside-Rd at a dose of 1 or 5 mg kg(-1) daily for 30 days to 10-month-old SAM significantly increased GSH, but decreased GSSG, resulting in elevation of the GSH/GSSG ratio. In addition, ginsenoside-Rd increased the activity of glutathione peroxidase (GSH-Px) and glutathione reductase that were both significantly lower in old SAM than in young SAM. This suggests that ginsenoside-Rd could play a crucial role in enhancing the defence system through regulation of the GSH/GSSG redox status. Moreover, decreases in the superoxide dismutase (SOD) and catalase activity in old SAM compared with young SAM were also revealed, indicating that the aging process resulted in suppression of the antioxidative defence system. However, ginsenoside-Rd did not affect SOD and catalase activity. As catalase is localized in peroxisome granules and GSH-Px is present in the cytoplasm and mitochondrial matrix, the site of ginsenoside-Rd action may be the cytoplasm and mitochondrial matrix. Furthermore, the serum and liver malondialdehyde levels, indicators of lipid peroxidation, were elevated with aging, while ginsenoside-Rd inhibited lipid peroxidation. This study indicates that the aging process leads to suppression of the antioxidative defence system and accumulation of lipid peroxidation products, while ginsenoside-Rd attenuates the oxidative damage, which may be responsible for the intervention of GSH/GSSG redox status.     

A novel biologically active seleno-organic compound--I. Glutathione peroxidase-like activity in vitro and antioxidant capacity of PZ 51 (Ebselen)

a synthetic seleno-organic compound, 2-phenyl-1,2-benzoisoselenazol-3(2H)-one (PZ 51), exhibits GSH peroxidase-like activity in vitro, in contrast to its sulfur analog, PZ 25. In addition, PZ 51 behaves as an antioxidant shown by a temporary protection of rat liver microsomes against ascorbate/ADP-Fe-induced lipid peroxidation, an effect also elicited by PZ 25 but to a smaller extent. This protection against lipid peroxidation is independent of GSH and of P-450 monooxygenase activity.     

Neuroprotective effect of liquiritin against focal cerebral ischemia/reperfusion in mice via its antioxidant and antiapoptosis properties

Our present study was conducted to investigate whether liquiritin (7-hydroxy-2-[4-[3,4,5-trihydroxy-6-(hydroxymethyl) oxan-2-yl] oxyphenyl]-chroman-4-one, 1), an active component of Glycyrrhiza uralensis Fisch., exerts a neuroprotective effect against focal cerebral ischemia/reperfusion (I/R) in male Institute of Cancer Research (ICR) mice. On the establishment of mice with middle cerebral artery occlusion (MCAO) for 2 h and reperfusion for 22 h, liquiritin at the doses of 40, 20, and 10 mg/kg was administered before MCAO once a day intragastrically for a subsequent 3 days. Neurological deficits and infarct volume were measured, respectively. The levels of malondialdehyde (MDA) and carbonyl, activities of superoxide anion (SOD), catalase (CAT) and glutathion peroxidase (GSH-Px) and reduced glutathione/oxidized disulfide (GSH/GSSG) ratio in brain were estimated spectrophotometrically. 8-Hydroxy-2′-deoxyguanosine (8-OHdG) and terminal deoxynucleotidyl transferase-mediated DuTP-biotin nick end labeling (TUNEL)-positive cells were detected by immunohistochemical analysis. Our results showed that the neurological deficits, infarct volume, and the levels of MDA and carbonyl decreased, the ratio of GSH/GSSG and the activities of SOD, CAT, and GSH-Px were compensatorily up-regulated, and 8-OHdG and TUNEL-positive cells decreased after 22 h of reperfusion in liquiritin-treated groups. These findings suggest that liquiritin might be a potential agent against cerebral I/R injury in mice by its antioxidant and antiapoptosis properties.     

异丙酚麻醉对腹部手术患者抗氧化保护作用

目的:研究异丙酚的抗氧化保护作用.方法:32例上腹部手术患者随机分为两组,治疗组静脉推注异丙酚10 mg•kg 1•h 1,对照组吸入3%安氟醚维持麻醉,分别于麻醉前,麻醉1,2,3 h抽取血样测定6 磷酸葡萄糖脱氢酶(G 6 PD)的活性、还原型谷胱甘肽(GSH)和氧化型谷胱甘肽(GSSG)的含量,并计算GSH/GSSG比值.结果:治疗组G 6 PD活性轻度升高,GSH/GSSG比值轻度降低;对照组G 6 PD活性显著升高,GSH/GSSG比值显著降低;两组比较差异有极显著性(P＜0.01).结论:异丙酚具有显著的抗氧化活性,是一种既具有麻醉效能又具有抗氧化保护作用的麻醉药物.     

Neuroprotective effect of liquiritin against focal cerebral ischemia/reperfusion in mice via its antioxidant and antiapoptosis properties

Our present study was conducted to investigate whether liquiritin (7-hydroxy-2-[4-[3,4,5-trihydroxy-6-(hydroxymethyl) oxan-2-yl] oxyphenyl]-chroman-4-one, 1), an active component of Glycyrrhiza uralensis Fisch., exerts a neuroprotective effect against focal cerebral ischemia/reperfusion (I/R) in male Institute of Cancer Research (ICR) mice. On the establishment of mice with middle cerebral artery occlusion (MCAO) for 2 h and reperfusion for 22 h, liquiritin at the doses of 40, 20, and 10 mg/kg was administered before MCAO once a day intragastrically for a subsequent 3 days. Neurological deficits and infarct volume were measured, respectively. The levels of malondialdehyde (MDA) and carbonyl, activities of superoxide anion (SOD), catalase (CAT) and glutathion peroxidase (GSH-Px) and reduced glutathione/oxidized disulfide (GSH/GSSG) ratio in brain were estimated spectrophotometrically. 8-Hydroxy-2'-deoxyguanosine (8-OHdG) and terminal deoxynucleotidyl transferase-mediated DuTP-biotin nick end labeling (TUNEL)-positive cells were detected by immunohistochemical analysis. Our results showed that the neurological deficits, infarct volume, and the levels of MDA and carbonyl decreased, the ratio of GSH/GSSG and the activities of SOD, CAT, and GSH-Px were compensatorily up-regulated, and 8-OHdG and TUNEL-positive cells decreased after 22 h of reperfusion in liquiritin-treated groups. These findings suggest that liquiritin might be a potential agent against cerebral I/R injury in mice by its antioxidant and antiapoptosis properties.     

N-acetylcysteine and glutathione-dependent protective effect of PZ51 (Ebselen) against diquat-induced cytotoxicity in isolated hepatocytes

The glutathione peroxidase (GSH-Px)-like reduction of H2O2 by the selenoorganic compound 2-phenyl-1,2-benzoisoselenazol-3(H)-one (PZ51: Ebselen) was studied using glutathione (GSH) and the therapeutic agent N-acetylcysteine (NAC) to provide reducing equivalents. In a purely chemical system containing H2O2 and in an enzymatic system of glucose/glucose oxidase-generated H2O2 Ebselen alone did not reduce H2O2. Ebselen in combination with either GSH (1 mM) or NAC (1 mM) was capable of reducing H2O2 in both systems. In these non-cellular systems GSH was a more effective source of reducing equivalents than NAC. The GSH-Px-like activity of Ebselen was further investigated in a cellular system. The redox-cycling bipyridylium compound diquat generates active oxygen species, depletes intracellular glutathione, and is cytotoxic in isolated hepatocytes pretreated with the glutathione reductase inhibitor 1,3-bis(Z-chloro-ethyl)-1-nitrosourea (BCNU). Ebselen alone did not ameliorate diquat cytotoxicity, but in combination with either GSH (1 mM) or NAC (1 mM) it produced a significant delay in diquat-induced cytotoxicity. Further additions of either GSH (0.5 mM) or NAC (0.5 mM) at 30 min intervals provided significantly more protection against diquat-induced cytotoxicity and intracellular GSH depletion than the single 1 mM addition. Thus, the combination of Ebselen and NAC may provide an effective antidote in cases of overexposure to bipyridylium herbicides, such as diquat and paraquat.     

葛根素对青光眼视网膜氧化应激损伤的保护作用

【摘要】目的研究葛根素对青光眼视网膜氧化应激损伤的保护作用。方法24只健康雄性成年家兔随机分为对照组、模型组及治疗组,每组8只。将2.5%羟丙基甲基纤维素溶液0.2mL注入家兔前房内,制作模型组及治疗组兔青光眼模型,治疗组家兔腹腔注射葛根素10mL/kg,每日1次,共21d。模型建立后第4周,测定视网膜抗氧化酶超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）和过氧化氢酶（CAT）活性,检测抗氧化物质谷胱甘肽含量及氧化还原状态和丙二醛（MDA）、一氧化氮（NO）含量。结果模型组视网膜抗氧化酶SOD、GSH-Px和CAT活性、还原型谷胱甘肽（GSH）和GSH/氧化型谷胱甘肽（GSSG）均低于对照组,GSSG、MDA和NO含量高于对照组;治疗组视网膜上述抗氧化酶活性、GSH含量、GSH/GSSG均高于模型组,但低于对照组;GSSG、MDA和NO含量均低于模型组,GSSG含量与对照组差异无统计学意义。结论葛根素可以在一定程度上增强抗氧化酶活性及维持谷胱甘肽氧化还原状态,从而减轻青光眼视网膜的氧化应激损伤。      

Nickel-induced renal lipid peroxidation in different strains of mice: concurrence with nickel effect on antioxidant defense systems

Lipid peroxidation (LPO) and active oxygen-detoxifying enzymes, catalase (CAT), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD), as well as glutathione (GSH) and some related enzymes, glutathione-S-transferase (GST) and glutathione reductase (GSSG-R) were assayed in kidneys of BALB/cAnNCr (BALB/c), C3H/HeNCr-MTV- (C3H), B6C3F1, and C57BL/6NCr (C57BL) mice 3-48 h after a single intraperitoneal injection of 170 mumol nickel (II) acetate (NiAcet)/kg body wt. In control mice that received 340 mumol sodium acetate/kg, the levels of enzymes and GSH did not significantly vary in time but were different in various strains. The basal activities of CAT and SOD in in the controls were highest in BALB/c and lowest in C57BL mice (1.8:1.0 and 1.4:1.0 respectively) in contrast to that of GSH-Px which was highest in B6CF1 and lowest in BALB/c (1.3:1.0; P less than 0.05). The strain ranking of control concentrations of renal GSH was B6C3F1 greater than C3H greater than or equal to C57BL greater than BALB/c (2.8:2.4:2.3:1.0), and that of GSSG-R was C3H greater than or equal to BALB/c greater than B6C3F1 greater than or equal to C57BL [corrected] (1.5:1.4:1.1:1.0). The basal activity of renal GST in control mice was 25% lower in C3H than in any of the other 3 strains. The renal LPO levels in the control mice did not vary among strains. Nickel treatment transiently increased renal LPO levels in the control mice did not vary among strains. Nickel treatment transiently increased renal LPO in the BALB/c mice by 100%, in B6C3F1 by 30%, and in C57BL by 20% (P less than 0.05), with no significant effect in C3H mice. Thus, the magnitude of nickel-induced renal LPO was greatest in the strain that is lowest in GSH and GSH-Px, but not in CAT and SOD. Nickel effects on GSH and the enzymes were time-dependent and included transient inhibition or enhancement of different proportions with no apparent strain- and/or base level-related patterns, or concurrence with LPO. The results emphasize the importance of GSH and GSH-Px for preventing nickel-induced oxidative cell damage.     

Increase in hepatic mixed disulphide and glutathione disulphide levels elicited by paraquat

Paraquat (1 mM), when added to isolated haemoglobin-free perfused rat liver, leads to an increase of intracellular mixed disulphides from 1.3 μmole GSH equivalents per g wet weight in the controls to 2.5 μmole/g. This raises the proportion of mixed disulphides to total glutathione equivalents from about 0.2 at the beginning of the perfusion to about 0.4. The mixed disulphides are predominantly protein-bound, with low molecular weight compounds being quantitatively negligible.The content of intracellular glutathione disulphide (GSSG) is increased from 17 nmole/g in the controls to 38 nmole/g in the presence of paraquat. In addition, there is an increased rate of release of GSSG into the extracellular (biliary) space, reported previously.It is suggested that, in a reaction catalysed by thioltransferase(s), the rise in GSSG is correlated with the rise in mited disulphides (reaction 1). Occupancy of potential cellular mixed disulphide sites is about $\text{1}\text{2}$ in the controls, and rises to about $\text{2}\text{3}$ in the presence of paraquat.The ratio of cellular contents, NADPH/NADP⁺, is decreased from 5.1 in the controls to 2.3 in the presence of paraquat, while the sum of NADPH plus NADP⁺ remains unaltered.The perturbation in the glutathione status may be related to metabolic effects such as the stimulation of the pentose-phosphate pathway activity, and possibly also to the expression of toxic effects.     

Inhibition of the activity of glutathione peroxidase by tertiary-butylhydroperoxide in cultured Chinese hamster cells and the role of cellular glutathione in the recovery of the activity.

Treatment of Chinese hamster cells with 1 mM tertiary-butylhydroperoxide (t-BuOOH) for 1 h markedly inhibited the activity of glutathione peroxidase (GSH-Px). However, the activity returned to pre-inhibition levels within 1-2 h with post-treatment incubation. The inhibition of the activity of GSH-Px by t-BuOOH was enhanced by depletion of levels of cellular GSH with the addition of L-buthionine sulfoximine (BSO) or diethylmaleate (DEM) and no subsequent recovery of the activity was observed for at least 4 h. The ratio of levels of GSSG to total GSH increased as a result of treatment with 1 mM t-BuOOH or diamide for 1 h but not as a result of treatment with hydrogen peroxide. After treatment with t-BuOOH, the level of GSH rapidly increased to more than twice the control level during 15-40 min of post-treatment incubation. Depletion of GSH by DEM after treatment with t-BuOOH reduced the rate of the recovery of the activity of GSH-Px, suggesting a role of cellular GSH in the recovery. However, the decrease in the rate of the recovery caused by DEM was small and in no way equivalent to the extent of depletion of GSH, suggesting that the rapid increase in the level of GSH after treatment with t-BuOOH is not closely related to the rapid recovery of the activity of GSH-Px.     

Taurodeoxycholate-induced intestinal injury is modulated by oxidative stress-dependent pre-conditioning like mechanisms

Mechanisms by which hydrophobic bile salts cause tissue changes below their critical micellar concentration (CMC, 1-2mM) and above (4-8mM) remain poorly understood. In this study, rat colonic mucosa was exposed to different concentrations of taurodeoxycholate (TDC), t-butyl-hydroperoxide (t-BH) or glutathione ester with or without pre-incubation with 2mM TDC. Exposure to 2mM TDC was associated with 10% higher tissue levels of total glutathione (GSH, basal values: 33.7+/-3.3 nmol/mg prot). With TDC 8mM, GSH decreased to 16.4+/-2.3 nmol/mg prot (P<0.05), oxidized glutathione (GSSG) increased by 60% (P<0.05), glutathione peroxidase (GSH-Px) and reductase activities were threefold increased, protein carbonyls fourfold increased, protein sulfhydrils decreased by 78%, lactate dehydrogenase (LDH) and GSSG release in the incubation medium were sixfold higher. In 2mM TDC pre-treated tissues, the subsequent incubation with 8mM TDC induced a lower loss of tissue GSH, and a lower release of LDH and GSSG. Pre-incubation with 2mM TDC partly protected against t-BH toxicity, while glutathione ester protected against 8mM TDC toxicity. In conclusion, TDC exposure causes opposite effects depending on CMC: induction of antioxidant protective systems including glutathione system (pre-conditioning effect) was observed with TDC below CMC, oxidative damages pointing to decreased mucosal detoxification potential with above CMC.     

Glutathione redox status of control and cadmium oxide-exposed rat lungs during oxidant stress

Activities of enzymes responsible for the maintenance of reduced glutathione (GSH) levels have been shown in a previous study to be increased in rat lungs following a 3-h exposure to cadmium oxide aerosols at 5.0 mg/m3. In this study, the ability of the lung to maintain levels of GSH during challenge with tert-butyl hydroperoxide (tBuOOH) was evaluated in isolated perfused lungs from control and cadmium oxide-exposed rats. Changes in glutathione redox status were indicated by measurements of nonprotein sulfhydryls (NPSH), total glutathione (1/2 GSH + GSSG), and glutathione disulfide (GSSG) in liquid nitrogen freeze-clamped lungs after 3-min infusions with 0-0.6 mM tBuOOH. In control and cadmium oxide-exposed lungs, levels of 1/2 GSH + GSSG remained constant over the range of 0-0.6 mM tBuOOH, indicating that no loss of glutathione from the system had occurred. In experiments with control lungs, levels of NPSH fell from 8.04 +/- 0.22 to 3.09 +/- 0.40 mumol/g dry weight when tBuOOH concentrations were increased from 0 to 0.6 mM (n = 20-23). In cadmium oxide-exposed lungs, NPSH levels also decreased proportionally to increases in GSSG. However, at concentrations of 0.075 and 0.15 mM tBuOOH, significantly smaller decreases in NPSH levels were observed in cadmium oxide-exposed lungs compared with controls. This protection against the GSH-depleting effects of tBuOOH might be explained by increased tissue levels of GSH-related enzymes.     

Metabolism of haloforms to carbon monoxide--III. Studies on the mechanism of the reaction

Haloforms are metabolized to carbon monoxide by hepatic microsomal mixed function oxidases and this reaction is markedly stimulated by sulfhydryl compounds. Maximal stimulation occurred at 0.5 mM glutathione (GSH). Formation of 1 mole of carbon monoxide (CO) resulted in the disappearance of 2 moles of GSH and the production of 1 mole of oxidized glutathione (GSSG). Incubation of ¹³CHBr₃, or ¹²CHBr₃ in the presence of ¹⁸O₂, resulted in the formation of similarly enriched ¹³CO or C¹⁸O respectively. Furthermore, a primary isotope effect was observed when CDBr₃ served as the substrate. Dibromocarbonyl is an intermediate in the reaction since 2-oxothiazolidine-4-carboxylic acid (OTZ) was detected when CHBr₃ was incubated in the presence of cysteine. In addition, when ¹³CHBr₃ was used, a similarly enriched [¹³C]OTZ was formed. Based on these observations, the following mechanism for the conversion of haloforms to CO is proposed: CHX → COHX₃ → X₂CO; X₂CO + GSH → GS(C = O)X; GS(C = O)X + GSH → GSSG + :C = O.     

Effect of exercise training on antioxidant system in brain regions of rat

The purpose of this investigation was to determine whether any alterations in antioxidant enzyme activities and levels of glutathione (GSH) in brain regions occurred following exercise training. Sprague-Dawley rats were given exercise training on a treadmill for 7.5 weeks and sacrificed 18 h after the last exercise along with the sedentary control rats. Different brain regions—cerebral cortex (CC), brainstem (BS), corpus striatum (CS), and hippocampus (H)—were isolated; GSH, oxidized glutathione (GSSG), Superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were determined. The exercise training increased SOD activity significantly (130% of sedentary control) in BS and in CS. SOD activity in H was the lowest of all four brain regions. Different brain regions showed GSH-Px activity in decreasing order for CS < BS < CC < H. GSH levels were 43% less in BS than CC and CS. The ratio of GSH/ GSSG significantly increased from 6.8 to 8.3 in CC, and from 9.4 to 13.5 in BS as a result of exercise training. Different brain regions contained different activities of antioxidant enzymes, as well as GSH and GSSG levels, which were preferentially altered as a result of exercise training to cope with oxidative stress.