Erythrocyte glutathione levels in lithium-induced hypothyroidism

BACKGROUND AND OBJECTIVE: Lithium, widely used in the prophylaxis of psychiatric patients with bipolar affective disorders, is well known to induce thyroid alterations. However, a possible metabolic linkage between blood thyroxine levels and its regulatory function on erythrocyte glutathione concentration has not yet been evaluated in lithium-treated patients. The aim of this study was to assess the antioxidative capacity of erythrocytes in lithium-induced hypothyroidism before and after thyroxine replacement. PATIENTS AND METHODS: Thyroid ultrasound with clinical and laboratory evaluation of thyroid function and thyroid-stimulating hormone assay were performed prior to and at 6-month intervals during lithium prophylaxis in 76 patients with bipolar affective disorders. The daily lithium dosage was adjusted to 600-1200 mg and the mean duration of treatment was 2.2 +/- 0.4 years. Final assessment revealed that 12 patients had evidence of primary hypothyroidism, and these were assigned as the test group. Lithium prophylaxis was supplemented with thyroxine at a dosage of 100 mg/day within 6 months after thyroid dysfunction was diagnosed. Red blood cell superoxide dismutase activities and the glutathione contents were measured before and after thyroxine replacement. In order to assess the effect of long-term lithium administration on red blood cell glutathione and superoxide dismutase levels, 12 patients who had not developed hypothyroidism during the follow-up period were selected for the lithium-treated euthyroid group. Mann Whitney U-test and Wilcoxon rank sum W-test were used for comparison of data. RESULTS: A comparison of the lithium-treated test group with healthy volunteers and their own values after thyroxine replacement revealed a significant decrease in red blood cell glutathione concentrations (p = 0.000) in the hypothyroid state. However, no clinically significant changes were observed in red blood cell superoxide dismutase activities of the test group. A statistical survey also demonstrated that there was no significant difference in the thyroid-stimulating hormone values as well as the red blood cell glutathione contents or superoxide dismutase activities between healthy controls and lithium-treated euthyroid subjects. CONCLUSIONS: It is most likely that lithium primarily inhibited hormone production in the thyroid and that this led to a compensatory increase in thyroid-stimulating hormone secretion with a significant decrease in the red blood cell glutathione content. While the red blood cell glutathione content of hypothyroid patients was reduced to 40% of the post-thyroxine level, unchanged superoxide dismutase activity might render the erythrocytes vulnerable to oxidative stress and ultimately haemolysis. Thyroxine replacement during lithium prophylaxis of psychiatric patients is advisable in order to prevent subclinical hypothyroidism and related defects of erythrocyte antioxidant capacity.     

Salicylate and liver glutathione

The effects of sodium salicylate, benzoate, gentisate, m- and p-hydroxybenzoates, antipyrine, and 2:4-dinitrophenol on the reduced glutathione concentration of rat liver were determined. An increase in concentration in the male, but not in the female, rats was found after salicylate, benzoate and gentisate; after antipyrine, a comparable increase occurred in the females only. There is a relation, in these compounds, between capacity to increase the concentration of reduced glutathione in the rat liver in one sex and therapeutic activity in rheumatic fever.     

Erythrocyte and plasma lipids in liver diseases.

Forty-four patients aged between 12 and 64 years comprising 16 hepatitis (group 1); 12 cirrhosis (group 2); 16 primary liver cell carcinoma (group 3) and 18 normal controls were studied. In hepatitis, plasma total cholesterol and total cholesterol/phospholipid ratio were significantly reduced, while the changes in red cell cholesterol and phospholipid and plasma phospholipid were not. The blood glucose was significantly reduced. The plasma total cholesterol/phospholipid ratio was positively correlated with the plasma total bilirubin. In cirrhosis patients, red cell total cholesterol and ratio to phospholipid were significantly increased and the plasma cholesterol reduced with no significant changes in red cell and plasma phospholipids. The plasma total cholesterol/phospholipid ratio was reduced while the corresponding ratio in red cells was increased. Both total cholesterol and the ratio to phospholipid in red cells were negatively correlated with albumin and positively correlated with the plasma total bilirubin. In primary liver cell carcinoma, the plasma and red cholesterol and their ratio in the red cell were significantly increased while the ratio in plasma was not. The serum albumin levels were reduced while the liver enzymes and total bilirubin were raised in all patient groups. Our results suggest a possible relationship between liver function and cholesterol deposition in red cells in liver disease.     

Changes in kinetic characteristics of sheep liver microsomes following induction and dietary stress

The effects of dietary stress and polychlorinated biphenyls (PCB) on the kinetic characteristics of ovine liver microsomal oxidases were studied. Curvilinear Lineweaver-Burke kinetic plots were obtained for the O-deethylation of p-nitrophenetole. Increasing linearity of the plots resulted from microsomal induction due to both dietary stress and PCB exposure. This suggests two enzyme systems with different magnitudes of induction catalyzing the same reaction.     

中国汉族196人红细胞谷胱甘肽硫转移酶的活性分布

目的采用分光光度法测定人RBC谷胱甘肽硫转移酶(GSTs)活性,研究196名(男101名,女95名)健康汉族人的GSTs活性分布。方法制备人RBC裂解后,参照Habdous等的方法测定GSTs活性。反应体系如下:0.1 mol.L-1磷酸钠缓冲液(pH 6.5)2.6 mL,RBC裂解液0.1 mL,30 mmol.L-1谷胱甘肽0.1 mL,10 mmol.L-11-氯-2,4-二硝基苯(CDNB)0.1 mL。结果方法的日内和日间精密度均<10%。RBC GSTs活性呈正偏态分布,196例健康汉族人的平均GSTs活性为(3.89±0.96)U.(gHb)-1。不同年龄组平均GSTs活性无差异,而女性的平均GSTs活性比男性的平均值高,但并不显著。结论确定了可供临床参考的健康汉族人GSTs活性分布范围。     

Erythrocyte glutathione, glutathione peroxidase, superoxide dismutase and serum lipid peroxidation in epileptic children with valproate and carbamazepine monotherapy

This prospective study was carried out to determine changes in the antioxidant system in epileptic children receiving long term antiepileptic drugs (AEDs). Levels of erythrocyte glutathione (GSH), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and serum lipid peroxidation were determined in 25 healthy children and 30 epileptic children who had not yet received AEDs. Sixteen patients were treated with valproic acid (VPA) and 14 with carbamazepine (CBZ); 13 months later these parameters were retested. The results showed that SOD and lipid peroxidation levels were increased but the GSH-Px levels were decreased in epileptic children on VPA therapy compared with the control group and the results before treatment. No significant differences of these parameters were found in epileptic children on CBZ therapy compared with the control group and the results before treatment, except that lipid peroxidation level was slightly higher in epileptic patients before treatment. We conclude that antioxidant systems in epileptic children on CBZ therapy are better regulated in comparison with epileptic children on VPA therapy.     

Inhibition of glutathione peroxidase and glutathione transferase in mouse liver by misonidazole

The mechanisms of toxicity and sensitization by the radiosensitizer misonidazole [1-(2-nitro-1-imidazolyl)-3-methoxy-2-propanol] are not well understood. We report here on the inhibition of total glutathione peroxidase (GSHPx), selenium-dependent glutathione peroxidase (selenium-GSHPx) and glutathione transferase (GSHTx) activities by misonidazole. Mouse liver cytosol GSHPx and selenium-GSHPx were inhibited in vitro with 0.5 mM misonidazole. On administration of the drug intraperitoneally (800 mg/kg) to mice, it was found that GSHPx, selenium-GSHPx, and GSHTx were inhibited in homogenate, cytosol, and microsomal fractions of mouse liver. GSHPx was depressed in all fractions up to 60-70% of control values, with maximum depression occurring in the cytosol and homogenate fractions in less than 2 hr. Recovery of activity was slower in the microsomes. In general, the pattern of depression of selenium-GSHPx was parallel to that of GSHPx except in microsomes, where GSHPx is minimal. Quantitatively, selenium-GSHPx was least affected. GSHTx was inhibited 70-80% of control values in cytosol and homogenate with recovery by 24 hr, whereas a second period of depression occurred at 24 hr in the microsomes. The inhibition of peroxide-metabolizing enzymes may lead to elevation of intracellular peroxide levels, contributing to the radiosensitizing effect and/or toxicity of misonidazole.     

Selenium- and non-selenium-dependent glutathione peroxidases in mouse liver

About 70% of the activity of mouse liver peroxidase utilizing H₂O₂ as well as t-butyl hydroperoxide as substrates is the selenium-dependent glutathione peroxidase. With cumene hydro-peroxide as substrate the sum of non-Se-dependent and Se-dependent enzymes has been measured. Se-deficient mice have about 30% of the total activity of Se-adequate mice, with cumene hydroperoxide as substrate. Glutathione S-transferase activity was not affected by Se deficiency. Fractionation on DEAE-Sepharose and subsequently on CM-Sepharose showed at least four transferase activities, and besides the Se-GSH-Px, two non-Se-GSH-Px activities detectable with cumene hydroperoxide that did not coincide with any of the transferases. For the major activity the apparent Michaelis constants at 1 mmole/l GSH were found to be 0.07mM for linoleic acid hydroperoxide, 10 mM for t-butyl hydro-peroxide, and 0.11 mM for cumene hydroperoxide. The ratios of the apparent maximum velocities of these hydroperoxides were 1 (linoleic): 100 (butyl) :3000 (cumene). This has the practical consequence that the cumene hydroperoxide-supported reaction is extremely favored under standard assay conditions. The results suggest that data based on only a single organic peroxide and hydrogen peroxide can lead to invalid conclusions as to the nature of the enzymes that are capable of catalyzing GSH-linked reduction of hydroperoxides.Subcellular fractionation showed that cytosol, mitochondria and microsomes contain both types of peroxidase, although in different proportions. The distribution as well as the kinetic properties suggest the Se-GSH-Px is exceedingly faster in reducing organic, including fatty acid, hydroperoxides.     

Effect of butylated hydroxytoluene on glutathione S-transferase and glutathione peroxidase activities in rat liver

When rats were fed a diet containing 0.4% (w/w) butylated hydroxytoluene (BHT), glutathione (GSH) S-transferase activity towards 1-chloro-2,4-dinitrobenzene (CDNB) increased approximately 3-fold in the liver. Immunotitration studies using the antibodies raised against rat liver GSH S-transferase B and GSH S-transferase A and C indicated that the increase in GSH S-transferase activity was probably due to de novo protein synthesis. Since some forms of rat liver GSH S-transferases express GSH peroxidase II activity, a concomitant increase in GSH peroxidase II was expected. However, GSH peroxidase II activity in the liver of BHT-treated rats remained unchanged. Gel filtration of supernatant fractions from livers of control and BHT-treated rats, followed by isoelectric focusing, indicated that BHT induced the activity of hepatic GSH S-transferases, without any apparent effect on GSH peroxidase II activity.     

Effect of monensin on liver glutathione, glutathione S-transferase and monooxygenases in rats.

Monensin administered ip to male rats at a dosage of 2.5 mg/kg/d for 3 consecutive days did not change the liver levels of glutathione, but depressed significantly the amount of cytochrome P-450 and the activities of aniline hydroxylase and a cytosolic CDNB-specific glutathione S-transferase. There was a marked decrease in the aminopyrine N-demethylase activity and a significant increase in the pentobarbital sleeping time in rats treated with monensin. In contrast, no change in these parameters was found 2 h after a single ip dose (7.5 mg/kg) of monensin. The results suggest that monensin-induced inhibition of the liver cytosolic glutathione S-transferase and microsomal monooxygenases is non-specific.     

Enzymatic changes in rat liver associated with low and high doses of a peroxisome proliferator

The activities of a number of lipid-metabolizing and subcellular marker enzymes were measured in total homogenates and subcellular fractions prepared from the livers of male rats fed diets containing 0.05, 0.1, 0.3, and 0.5% of the hypolipidemic drug tiadenol, resulting in mean drug intake of 45, 90, 330, and 530 mg/day/kg body wt, respectively. In the total homogenates, a massive induction of palmitoyl-CoA hydrolase and peroxisomal palmitoyl-CoA oxidation accompanied by increased free CoASH and long-chain acyl-CoA content was observed at the highest dose levels whereas little change occurred up to 90 mg/day/kg/body wt. The palmitoyl-CoA synthetase activity increased slightly up to 90 mg/day/kg body wt, but higher doses resulted in decreased enzyme activity. Catalase activity increased with the dose to be elevated by a factor of approximately 1.6 at 330 mg/day/kg, whereas the activities of urate oxidase decreased. The specific activities of palmitoyl-CoA hydrolase and peroxisomal palmitoyl-CoA oxidation increased in all fractions, but most markedly in the cytosol. The changes in the activities and the distribution of subcellular marker enzymes and the increase of the peroxisome-associated polypeptide (PPA-80) are in keeping with a peroxisome proliferating effect resulting in formation of premature organelles with altered properties. Since high doses of many hypolipidemic drugs produce hepatic tumors and peroxisomal proliferation in rodents and since no increase in peroxisomes is found in human liver after therapeutic use of lower doses, the dose-response relationship is of interest for the evaluation of the toxicology of this class of agents.     

Different effects of nine clausenamide ennatiomers on liver glutathione biosynthesis and glutathione S-transferase activity in mice

AIM: To study the effects of nine synthetic clausenamide with different stereo structures on liver glutathione (GSH) biosynthesis and glutathione S-transferase (GST) activity in mice. METHODS: The nine test compounds were racemic mixtures and their ennatiomers of clausenamide, neoclausenamide and epineoclausenamide. Mice were administered clausenamide 250 mg/kg once daily for 3 consecutive days, ig, and were killed 24 h after the last dosing. The mouse liver cytosol GSH and GST were determined with related biochemical methods. RESULTS: Nine clausenamides exhibited different effects on liver GSH and GST. Of nine clausenamides, only (+) and (+/-)clausenamide markedly increased liver cytosol GSH content. The mechanism of increasing liver GSH content of (+)clausenamide is mainly due to stimulating the key limiting enzyme gamma-glutamylcysteine synthetase (gamma-GCS) activity for GSH biosynthesis. The other test clausenamides had no such effect on liver GSH. All of the nine clausenamides induced a significant increase of GST activity. CONCLUSION: The effects of clausenamide ennatiomers on liver GST and GSH varied with the alterations of their spatial structures. (+)Clausenamide stimulated liver GSH biosynthesis through enhancing gamma-GCS activity.     

Ascorbic acid, dehydroascorbic acid and glutathione in liver disease

Controlled studies were conducted to find out the plasma values of ascorbic acid, dehydroascorbic acid (DHA), urinary excretion of ascorbic acid and blood levels of glutathione in patients with viral hepatitis, alcoholic hepatitis, cirrhosis of liver and carcinoma of liver. Leucocyte ascorbic acid and DHA/AA index were also determined in order to assess the ascorbic acid status of these patients. It was observed that the plasma and leucocytes contents of ascorbic acid were significantly subnormal with markedly decreased urinary excretion in these patients. Decreased level of glutathione and significantly higher level of DHA reflect an over all reducing status of the body is markedly deranged in these conditions. Further it was observed that the DHA/AA ratios were significantly altered in these groups of patients.     

Inhibition of rat liver glutathione S-transferases by glutathione conjugates and corresponding L-cysteines and mercapturic acids

Glutathione S-transferases from rat liver were partially purified by ion exchange chromatography. Active peaks, tentatively identified as containing the 1-2, 2-2, 3-3, 3-4, 4-4 and 5-5 isoenzymes were kept for study. The glutathione conjugates, S-hexyl-, S-benzyl- and S-(2,4-dinitrophenyl) L-glutathione were tested as inhibitors of the enzymes. The 1-2, 2-2, 3-3 and 3-4 fractions were inhibited to similar extents by these conjugates. For all enzymes the hexyl conjugate at 0.1 mM concentration was strongly inhibitory, the benzyl conjugate moderately so and the dinitrophenyl compound was only weakly inhibitory. In contrast, the epoxide conjugating activity in the 4-4 and 5-5 peak was barely affected by the substituted glutathiones at 0.1 mM concentrations. Studies on a purified ligandin (isoenzyme 1-2) from rat liver showed that further metabolism of the glutathione conjugates, to the corresponding cysteines or mercapturic acids, resulted in products with inhibitory properties approximately three orders of magnitude less potent than those of the parent S-substituted glutathiones.     

Subcellular distribution of glutathione S-transferase in Chinese fetal liver

Subcellular fractions were isolated from Chinese fetal liver at 4-8 months of age for the determination of glutathione S-transferase (GST). Using 1-chloro-2,4-dinitrobenzene (CDNB) as substrate, GST activity was found to be 66 +/- 34 nmol/(min.mg protein), mainly in the cytosol. The GST activities were detected principally in microsomes and their values were 66 +/- 31 and 144 +/- 83 nmol/(min.mg protein), respectively, when assayed with p-nitrobenzyl chloride (PNB) and ethacrynic acid (EA) as substrates. There were no age and sex-related differences in GST activities for any of the substrates studied during fetal development. The Km values of GST for CDNB, PNB and EA were 1112, 1039 and 205 mumol/L, respectively. The conjugation of GST may play an important role in fetal hepatic metabolism of toxic electrophiles.     

Formation of ortho-phenylphenol glutathione conjugates in the rat liver

1. The radioactive materials(s) derived from 14C-ortho-phenylphenol (OPP) became bound irreversibly to hepatic microsomal macromolecules in an NADPH-generating system, and the binding was inhibited by cysteine and glutathione (GSH). 2. A water-soluble phenyl-hydroquinol (PHQ)-GSH conjugate was produced by non-enzymic reaction between phenyl-p-benzoquinone and GSH in alcoholic aqueous solution, and the structure of the conjugate was confirmed by its 13C-n.m.r. spectrum. 3. When 14C-OPP and GSH were incubated in a microsomal NADPH-generating system, radioactive material derived from the aqueous phase of the incubation mixture was similar to the synthetic PHQ-GSH on t.l.c. 4. PHQ-GSH was excreted as a minor conjugate into the bile after oral administration of OPP (1000 mg/kg) to rats, the cumulative biliary excretion of the conjugate was about 4% of the dose in 6 h.     

Effects of ethanol on glutathione conjugation in rat liver and lung

The ability of ethanol to alter glutathione (GSH) conjugation and its dependence upon duration of administration were investigated in rats in correlation with lipid peroxidation and the induction of microsomal enzymes. Significant decreases in hepatic GSH and glutathione-S-transferase (GST) activity in both liver and lung were found in rats treated acutely with ethanol (4 g/kg body weight 6 hr prior to killing). These decreases were accompanied by an increased loss of both GSH and GST into the plasma and increased hepatic lipid peroxidation. On the other hand, there was a dose-dependent increase in hepatic GSH after chronic administration of ethanol in drinking water (5 and 10%) for 3 weeks. This increase in hepatic GSH may be due to increased synthesis of GSH in the liver. No significant induction of GST by chronic ethanol treatment was observed in either organ. Ethanol was compared with the well-known inducers phenobarbital and beta-naphthoflavone. Although there was some evidence of increases in lipid peroxidation and/or microsomal enzyme activity with the inducers, no simple link between these increases and the induction of GST activity was identified.     

腺苷蛋氨酸治疗药物性肝损害疗效与血液中GSH变化关系

目的　探讨应用腺苷蛋氨酸治疗药物性肝损害的疗效以及还原型谷胱甘肽 (GSH)对判断疗效的意义。方法　应用腺苷蛋氨酸对 1 4例因各种药物引起的肝损害给予治疗 ,连续静脉给药 80 0mg/d ,3周。动态观察治疗前后临床表现和各项肝功能指标及GSH水平变化情况。结果　治疗后精神、睡眠、食欲好转 ,乏力减轻 ,肝功能指标明显下降 ,GSH水平从治疗前的 6 2 1± 0 89mmol/L升至 7 40± 0 61mmol/L。结论　腺苷蛋氨酸对治疗药物性肝损害有明显疗效 ,能明显改善肝功能和抗氧化作用。检测血清GSH水平变化情况对判断肝损害的疗效及预后有重要意义