Ruth A. Sanders

Effects of α-lipoic acid on biomarkers of oxidative stress in streptozotocin-induced diabetic rats

Abstract Increased oxidative stress and impaired antioxidant defense mechanisms are important factors in the pathogenesis and progression of diabetes mellitus and other oxidant-related diseases. This study was designed to determine whether α-lipoic acid, which has been shown to have substantial antioxidant properties, when administered (10 mg/kg ip) once daily for 14 days to normal and diabetic female Sprague-Dawley rats would prevent diabetes-induced changes in biomarkers of oxidative stress in liver, kidney and heart. Serum glucose concentrations, aspartate aminotransferase activity, and glycated hemoglobin levels, which were increased in diabetes, were not significantly altered by α-lipoic acid treatment. Normal rats treated with a high dose of α-lipoic acid (50 mg/kg) survived but diabetic rats on similar treatment died during the course of the experiment. The activity of glutathione peroxidase was increased in livers of normal rats treated with α-lipoic acid, but decreased in diabetic rats after α-lipoic acid treatment. Hepatic catalase activity was decreased in both normal and diabetic rats after α-lipoic acid treatment. Concentrations of reduced glutathione and glutathione disulfide in liver were increased after α-lipoic acid treatment of normal rats, but were not altered in diabetics. In kidney, glutathione peroxidase activity was elevated in diabetic rats, and in both normal and diabetic animals after α-lipoic acid treatment. Superoxide dismutase activity in heart was decreased in diabetic rats but normalized after treatment with α-lipoic acid; other cardiac enzyme activities were not influenced by either diabetes or antioxidant treatment. These results suggest that after 14 days of treatment with an appropriate pharmacological dose, α-lipoic acid may reduce oxidative stress in STZ-induced diabetic rats, perhaps by modulating the thiol status of the cells.

Effects of coenzyme Q10 treatment on antioxidant pathways in normal and streptozotocin-induced diabetic rats

Coenzyme Q10 is an endogenous lipid soluble antioxidant. Because oxidant stress may exacerbate some complications of diabetes mellitus, this study investigated the effects of subacute treatment with exogenous coenzyme Q10 (10 mg/kg/day, i.p. for 14 days) on tissue antioxidant defenses in 30‐day streptozotocin‐induced diabetic Sprague‐Dawley rats. Liver, kidney, brain, and heart were assayed for degree of lipid peroxidation, reduced and oxidized glutathione contents, and activities of catalase, superoxide dismutase, glutathione peroxidase, and glutathione reductase. All tissues from diabetic animals exhibited increased oxidative stress and disturbances in antioxidant defense when compared with normal controls. Treatment with the lipophilic compound coenzyme Q10 reversed diabetic effects on hepatic glutathione peroxidase activity, on renal superoxide dismutase activity, on cardiac lipid peroxidation, and on oxidized glutathione concentration in brain. However, treatment with coenzyme Q10 also exacerbated the increase in cardiac catalase activity, which was already elevated by diabetes, further decreased hepatic glutathione reductase activity, augmented the increase in hepatic lipid peroxidation, and further increased glutathione peroxidase activity in the heart and brain of diabetic animals. Subacute dosing with coenzyme Q10 ameliorated some of the diabetes‐induced changes in oxidative stress. However, exacerbation of several diabetes‐related effects was also observed.

Effects of piperine on antioxidant pathways in tissues from normal and streptozotocin-induced diabetic rats

Using diabetes mellitus as a model of oxidative damage, this study investigated whether subacute treatment (10mg/kg/day, intraperitoneally for 14 days) with the compound piperine would protect against diabetes‐induced oxidative stress in 30‐day streptozotocin‐induced diabetic Sprague‐Dawley rats. Liver, kidney, brain, and heart were assayed for degree of lipid peroxidation, reduced and oxidized glutathione (GSH and GSSG, respectively) content, and activities of the free‐radical detoxifying enzymes catalase, superoxide dismutase, glutathione peroxidase, and glutathione reductase. Piperine treatment of normal rats enhanced hepatic GSSG concentration by 100% and decreased renal GSH concentration by 35% and renal glutathione reductase activity by 25% when compared to normal controls. All tissues from diabetic animals exhibited disturbances in antioxidant defense when compared with normal controls. Treatment with piperine reversed the diabetic effects on GSSG concentration in brain, on renal glutathione peroxidase and superoxide dismutase activities, and on cardiac glutathione reductase activity and lipid peroxidation. Piperine treatment did not reverse the effects of diabetes on hepatic GSH concentrations, lipid peroxidation, or glutathione peroxidase or catalase activities; on renal superoxide dismutase activity; or on cardiac glutathione peroxidase or catalase activities. These data indicate that subacute treatment with piperine for 14 days is only partially effective as an antioxidant therapy in diabetes.

Effects of Melatonin on Oxidative Stress in Streptozotocin-Induced Diabetic Rats

Oxidative stress plays an important role in diabetes and other oxygen-related diseases. Melatonin, a pineal hormone thought to be a scavenger of oxygen radicals and a potentially advantageous therapeutic agent in diseases having oxidative stress, was administered (10 mg/kg ip, in gum tragacanth to prolong its absorption, once a day for 4 successive days) to normal and 30-day streptozotocin-induced diabetic Sprague-Dawley rats, after which markers of oxidative stress were assessed in the liver, kidney, intestine, and spleen. Alanine and aspartate aminotransferase activities in serum, which were increased after diabetes, were not increased further by melatonin administration, indicating that there was no melatonin-related liver toxicity. Most melatonin-induced effects were seen in the liver, and very few in extrahepatic tissues. In livers of diabetic rats, reduced concentration of nitrite and increased lipid peroxidation were both restored to normal levels following treatment with melatonin. Hepatic glutathione peroxidase activity was not changed in diabetics, but was decreased after melatonin administration in both normal and diabetic animals. Total glutathione concentrations were significantly decreased in livers of all diabetics and were not normalized by melatonin treatment. Hepatic superoxide dis-mutase activity was elevated following melatonin dosing in normal rats, but dropped below normal levels in diabetic rats and was not restored by melatonin treatment. Glutathione 5-transferase activity was higher than normal in melatonin-dosed normal rat livers. These results suggest that after 4 days of administration, melatonin may enable various enzymes of the hepatic antioxidative defense system to better detoxify harmful oxygen radicals without producing overt toxicity in a disease such as diabetes.

The effect of long-term streptozotocin-induced diabetes on the hepatotoxicity of bromobenzene and carbon tetrachloride and hepatic biotransformation in rats

To exclude the possibility that changes in hepatotoxicity and biotransformation were induced by diabetogen administration, the influence of long-lasting experimental insulin-dependent diabetes on the activities of benzphetamine demethylase, styrene oxide hydrolase, and UDP-glucuronosyl-transferases toward 1-naphthol, diethylstilbestrol, estrone and testosterone, and glutathione S-transferases toward 1-chloro-2,4-dinitrobenzene, ethacrynic acid, and sulfobromophthalein was studied. Adult male Sprague-Dawley rats injected with 45 mg streptozotocin/kg rapidly developed the classical symptoms of diabetes which persisted throughout the 90-day test period. Ketonemia was detectable at 6 but not at either 35 or 90 days after streptozotocin administration. After acute challenge with bromobenzene or carbon tetrachloride (CCl4), aspartate and alanine aminotransferase activities in rats diabetic for 35 and 90 days were markedly higher than those in normal rats, suggesting that diabetes potentiated the hepatotoxicity of these chemicals. Administration of 25 microliters CCl4/kg, ip, to diabetic rats decreased enzyme activities toward benzphetamine, sulfobromophthalein, 1-chloro-2,4-dinitrobenzene, and 1-naphthol. In normal rats, a dose of 400 microliters CCl4/kg, ip, was required to cause similar changes in enzyme activities. Bromobenzene (500 microliters/kg, ip) elicited opposing responses in diabetic and normal rats in N-demethylase activity, in UDP-glucuronosyltransferase activity toward 1-naphthol, estrone, and testosterone, and in glutathione S-transferase activity toward 1-chloro-2,4-dinitrobenzene. Total cytochrome P450 concentrations were reduced by both induction of diabetes and hepatotoxicant challenge. Thus, chronic uncontrolled diabetes alters the response of hepatic xenobiotic biotransformation enzymes in a non-uniform, substrate-dependent manner, independent of initial diabetogen effects. The role of cytochrome P450j in potentiating CCl4 toxicity is discussed.

Effects of new antioxidant compounds PNU-104067F and PNU-74389G on antioxidant defense in normal and diabetic rats

Diabetes mellitus and its complications are associated with elevated oxidative stress, leading to much interest in antioxidant compounds as possible therapeutic agents. Two new classes of antioxidant compounds, the pyrrolopyrimidines and the 21‐aminosteroids, are known to inhibit lipid peroxidation and other biomolecular oxidation. We hypothesized that in the presence of excess oxidants or the impaired antioxidant defense seen in diabetes mellitus, administration of antioxidants such as these may reverse the effects of diabetes on antioxidant parameters. This study measured the effects of subchronic (14 day) treatment with a pyrrolopyrimidine (PNU‐104067F) or a 21‐aminosteroid (PNU‐74389G) in normal and diabetic Sprague‐Dawley rats. Activity levels of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, concentrations of oxidized and reduced glutathione, and lipid peroxidation were used as measures of antioxidant defense in liver, kidney, heart, and brain tissue. In normal rats, the only effect was a 43% increase in cardiac lipid peroxidation after treatment with PNU‐104067F. In diabetic rats, the only reversals of the effects of diabetes were a 30% decrease in hepatic glutathione peroxidase activity after PNU‐74389G treatment and a 33% increase in cardiac glutathione disulfide concentration after PNU‐104067F treatment. In contrast to these effects, increased cardiac glutathione peroxidase and catalase activities, increased brain glutathione peroxidase activity, increased hepatic lipid peroxidation, decreased hepatic glutathione content, and decreased hepatic catalase activity were seen in diabetic rats, reflecting an exacerbation of the effects of diabetes.

Effects of low-level light therapy on hepatic antioxidant defense in acute and chronic diabetic rats

Diabetes causes oxidative stress in the liver and other tissues prone to complications. Photobiomodulation by near infrared light (670 nm) has been shown to accelerate diabetic wound healing, improve recovery from oxidative injury in the kidney, and attenuate degeneration in retina and optic nerve. The present study tested the hypothesis that 670 nm photobiomodulation, a low‐level light therapy, would attenuate oxidative stress and enhance the antioxidant protection system in the liver of a model of type I diabetes. Male Wistar rats were made diabetic with streptozotocin (50 mg/kg, ip) then exposed to 670 nm light (9 J/cm2) once per day for 18 days (acute) or 14 weeks (chronic). Livers were harvested, flash frozen, and then assayed for markers of oxidative stress. Light treatment was ineffective as an antioxidant therapy in chronic diabetes, but light treatment for 18 days in acutely diabetic rats resulted in the normalization of hepatic glutathione reductase and superoxide dismutase activities and a significant increase in glutathione peroxidase and glutathione‐S transferase activities. The results of this study suggest that 670 nm photobiomodulation may reduce, at least in part, acute hepatic oxidative stress by enhancing the antioxidant defense system in the diabetic rat model.

Oxidative stress in rats after 60 days of hypergalactosemia or hyperglycemia.

Two of the models used in current diabetes research include the hypergalactosemic rat and the hyperglucosemic, streptozotocin-induced diabetic rat. Few studies, however, have examined the concurrence of these two models regarding the effects of elevated hexoses on biomarkers of oxidative stress. This study compared the activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase and the concentrations of glutathione, glutathione disulfide, and thiobarbituric acid reactants (as a measure of lipid peroxidation) in liver, kidney, and heart of Sprague-Dawley rats after 60 days of either a 50% galactose diet or insulin deficiency caused by streptozotocin injection. Most rats from both models developed bilateral cataracts. Blood glucose and glycosy-lated hemoglobin A1c concentrations were elevated in streptozotocin diabetic rats. Streptozotocin diabetic rats exhibited elevated activities of renal superoxide dismutase, cardiac catalase, and renal and cardiac glutathione peroxidase, as well as elevated hepatic lipid peroxidation. Insulin treatment of streptozotocin-induced diabetic rats normalized altered markers. In galactosemic rats, hepatic lipid peroxidation was increased whereas glutathione reductase activity was diminished. Glutathione levels in liver were decreased in diabetic rats but elevated in the galactosemic rats, whereas hepatic glutathione disulfide concentrations were decreased much more in diabetes than in galactosemia. Insulin treatment reversed/prevented all changes caused by streptozotocin-induced diabetes. Lack of concomitance in these data indicate that the 60-day galactose-fed rat is not experiencing the same oxidative stress as the streptozotocin diabetic rat, and that investigators must be cautious drawing conclusions regarding the concurrence of the effects of the two animal models on oxidative stress biomarkers.

Attenuation of TCDD-induced oxidative stress by 670 nm photobiomodulation in developmental chicken kidney.

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin (TCDD), a potent developmental teratogen inducing oxidative stress and sublethal changes in multiple organs, provokes developmental renal injuries. In this study, we investigated TCDD‐induced biochemical changes and the therapeutic efficacy of photobiomodulation (670 nm; 4 J/cm2) on oxidative stress in chicken kidneys during development. Eggs were injected once prior to incubation with TCDD (2 pg/g or 200 pg/g) or sunflower oil vehicle control. Half of the eggs in each dose group were then treated with red light once per day through embryonic day 20 (E20). Upon hatching at E21, the kidneys were collected and assayed for glutathione peroxidase, glutathione reductase, catalase, superoxide dimutase, and glutathione‐S‐transferase activities, as well as reduced glutathione and ATP levels, and lipid peroxidation. TCDD exposure alone suppressed the activity of the antioxidant enzymes, increased lipid peroxidation, and depleted available ATP. The biochemical indicators of oxidative and energy stress in the kidney were reversed by daily phototherapy, restoring ATP and glutathione contents and increasing antioxidant enzyme activities to control levels. Photobiomodulation also normalized the level of lipid peroxidation increased by TCDD exposure. The results of this study suggest that 670 nm photobiomodulation may be useful as a noninvasive treatment for renal injury resulting from chemically induced cellular oxidative and energy stress.

Minimal effect of acute experimental hepatitis induced by lipopolysaccharide/ d-galactosamine on biotransformation in rats

When administered with D-galactosamine, lipopolysaccharide endotoxins produce a good experimental animal model of hepatitis. This galactosamine plus endotoxin model has been used widely, but the acute effect of this fixed combination of two chemicals on hepatic and extrahepatic biotransformation has not been determined. Therefore, either 2 or 4 hr after a single intraperitoneal dose of 300 mg/kg galactosamine plus 30 micrograms/kg lipopolysaccharide was administered, serum, liver, kidney, intestine, and spleen were collected. Serum enzymes (alanine and aspartate aminotransferases, sorbitol dehydrogenase, and gamma-glutamyltranspeptidase) were elevated dramatically 2 and 4 hr after treatment. Cytochrome P450 monooxygenase activity toward benzo-[a]pyrene was increased in kidney 4 hr after treatment, whereas dealkylation of 7-methoxycoumarin or 7-ethoxyresorufin was unchanged in any tissue at either time point. An increase in UDP-glucuronosyltransferase activity toward 4-methylumbelliferone and 4-hydroxybiphenyl was noted in the intestine. Conjugation of 1-chloro-2,4-dinitrobenzene with glutathione was increased in intestine and spleen 2 hr after treatment. gamma-Glutamyltranspeptidase activity was unaltered in all tissues studied. Reduced glutathione concentrations were increased significantly by different amounts depending on which organs were studied 2 or 4 hr after treatment. These results indicate that galactosamine/lipopolysaccharide-induced liver injury is not accompanied by major effects on the examined biotransformation reactions.