High stability of blood parameters during mouse lifespan: sex-specific effects of every-other-day fasting.

Every-other-day fasting (EODF) is one type of caloric restriction that is proposed to have significant health benefits, including slowing aging-related processes. The present study evaluated multiple parameters of blood homeostasis comparing mice of different ages and mice on different diet regimes: ad libitum (AL) versus EODF. Hematological and classical biochemical parameters of blood were measured in young (6-month), middle-aged (12-month) and old (18-month) C57BL/6J mice of both sexes subjected either to EODF, or AL feeding. Middle-aged AL males showed a decrease in erythrocyte and total leucocyte counts and an increase in plasma alkaline phosphatase activity, whereas old animals showed a decrease in relative levels of lymphocytes and an increase in relative levels of neutrophils, a decrease in plasma lactate and an increase in total cholesterol levels, compared to young mice. AL-fed females demonstrated higher stability of blood parameters during aging than males did. The EODF regimen did not significantly affect hematological parameters in females but prevented a decline in total leukocyte count with age in males. In both sexes, EODF partially prevented age-associated changes in levels of plasma lactate and cholesterol and activity of alkaline phosphatase. Thus, during normal aging, mice showed a sex-dependent maintenance of blood homeostasis which was not significantly affected by EODF.

Middle aged turn point in parameters of oxidative stress and glucose catabolism in mouse cerebellum during lifespan: minor effects of every-other-day fasting.

The cerebellum is considered to develop aging markers more slowly than other parts of the brain. Intensification of free radical processes and compromised bioenergetics, critical hallmarks of normal brain aging, may be slowed down by caloric restriction. This study aimed to evaluate the intensity of oxidative stress and the enzymatic potential to utilize glucose via glycolysis or the pentose phosphate pathway (PPP) in the cerebellum of mice under ad libitum versus every-other-day fasting (EODF) feeding regimens. Levels of lipid peroxides, activities of antioxidant and key glycolytic and PPP enzymes were measured in young (6-month), middle-aged (12-month) and old (18-month) C57BL/6J mice. The cerebellum showed the most dramatic increase in lipid peroxide levels, antioxidant capacity and PPP key enzyme activities and the sharpest decline in the activities of key glycolytic enzymes under transition from young to middle age but these changes slowed when transiting from middle to old age. A decrease in the activity of the key glycolytic enzyme phosphofructokinase was accompanied by a concomitant increase in the activities of hexokinase and glucose-6-phosphate dehydrogenase (G6PDH), which may suggest that during normal cerebellar aging glucose metabolism shifts from glycolysis to the pentose phosphate pathway. The data indicate that intensification of free radical processes in the cerebellum occurred by middle age and that activation of the PPP together with increased antioxidant capacity can help to resist these changes into old age. However, the EODF regime did not significantly modulate or alleviate any of the metabolic processes studied in this analysis of the aging cerebellum.

Middle age as a turning point in mouse cerebral cortex energy and redox metabolism: Modulation by every-other-day fasting.

Normal brain aging is accompanied by intensification of free radical processes and compromised bioenergetics. Caloric restriction is expected to counteract these changes but the underlying protective mechanisms remain poorly understood. The present work aimed to investigate the intensity of oxidative stress and energy metabolism in the cerebral cortex comparing mice of different ages as well as comparing mice given one of two regimens of food availability: ad libitum versus every-other-day fasting (EODF). Levels of oxidative stress markers, ketone bodies, glycolytic intermediates, mitochondrial respiration, and activities of antioxidant and glycolytic enzymes were assessed in cortex from 6-, 12- and 18-month old C57BL/6J mice. The greatest increase in oxidative stress markers and the sharpest decline in key glycolytic enzyme activities was observed in mice upon the transition from young (6 months) to middle (12 months) age, with smaller changes occurring upon transition to old-age (18 months). Brain mitochondrial respiration showed no significant changes with age. A decrease in the activities of key glycolytic enzymes was accompanied by an increase in the activity of glucose-6-phosphate dehydrogenase suggesting that during normal brain aging glucose metabolism is altered to lower glycolytic activity and increase dependence on the pentose-phosphate pathway. Interestingly, levels of ketone bodies and antioxidant capacity showed a greater decrease in the brain cortex of females as compared with males. The EODF regimen further suppressed glycolytic enzyme activities in the cortex of old mice, and partially enhanced oxygen consumption and respiratory control in the cortex of middle aged and old males. Thus, in the mammalian cortex the major aging-induced metabolic changes are already seen in middle age and are slightly alleviated by an intermittent fasting mode of feeding.

Acute exposure to copper induces variable intensity of oxidative stress in goldfish tissues.

Copper is an essential element, but at high concentrations, it is toxic for living organisms. The present study investigated the responses of goldfish, Carassius auratus, to 96 h exposure to 30, 300, or 700 µg L-1 of copper II chloride (Cu2+). The content of protein carbonyls was higher in kidney (by 158%) after exposure to 700 mg L-1 copper, whereas in gills, liver, and brain, we observed lower content of protein carbonyls after exposure to copper compared with control values. Exposure to copper resulted in increased levels of lipid peroxides in gills (76%) and liver (95-110%) after exposure to 300 and 700 µg L-1 Cu2+. Low molecular mass thiols were depleted by 23-40% in liver and by 29-67% in kidney in response to copper treatment and can be used as biomarkers toxicity of copper. The activities of primary antioxidant enzymes, superoxide dismutase and catalase, were increased in liver as a result of Cu2+ exposure, whereas in kidney catalase activity was decreased. The activities of glutathione-related enzymes, glutathione peroxidase, glutathione-S-transferase, and glutathione reductase were decreased as a result of copper exposure, but glutathione reductase activity increased by 25-40% in liver. Taken together, these data show that exposure of fish to Cu2+ ions results in the development of low/high intensity oxidative stress reflected in enhanced activities of antioxidant and associated enzymes in different goldfish tissues.

Acute exposure to the penconazole-containing fungicide Topas partially augments antioxidant potential in goldfish tissues.

Penconazole is a systemic fungicide commonly used in agriculture as the commercial preparation Topas. Although triazole fungicides are widely found in the aquatic environment, little is known about their acute toxicity on fish. In this study we assessed the effects of short-term exposure to Topas on some parameters of homeostasis of reactive oxygen species (ROS), such as the levels of markers of oxidative stress and parameters of the antioxidant defense system of goldfish (Carassius auratus L.). Gills appeared to be the main target organ of Topas toxicity, showing the greatest number of parameters affected. Gills of Topas-treated fish showed a higher content of low (L-SH) and high (H-SH) molecular mass thiols and higher activities of superoxide dismutase (SOD), catalase, glutathione reductase (GR), glutathione-S-transferase (GST), and glucose-6-phosphate dehydrogenase (G6PDH) as well as reduced carbonyl protein content (CP), as compared with those in the control group. In the liver, goldfish exposure to 15-25mgL-1 Topas resulted in a higher L-SH and H-SH content, but lower CP levels and activity of GST. In kidney, Topas exposure resulted in higher activities of glutathione peroxidase (GPx) and G6PDH, but lower L-SH content and activity of GST. The results of this study indicate that acute goldfish exposure to the triazole fungicide Topas increased efficiency of the antioxidant system in fish gills, liver, and kidney. This could indicate the development of low intensity oxidative stress which up-regulates defense mechanisms responsible for protection of goldfish against deleterious ROS effects.

Oxidative stress responses in gills of goldfish, Carassius auratus, exposed to the metribuzin-containing herbicide Sencor.

Metribuzin belongs to the family of asymmetrical triazine compounds and is an active ingredient in many commercial herbicides including Sencor. Effects on goldfish (Carassius auratus L.) of exposure for 96h to 7.14, 35.7 or 71.4mgL(-1) Sencor 70 WG (corresponding to 5, 25 and 50mgL(-1) of metribuzin) were examined by evaluating oxidative stress markers and activities of antioxidant and associated enzymes in gills. Fish exposed to the lowest Sencor concentration (7.14mgL(-1)) showed a 94% increase in levels of protein carbonyls in gills as well as 45% and 144% increases in the activities of glutathione peroxidase and glutathione-S-transferase. Exposure to the highest Sencor concentration (71.4mgL(-1)) resulted in reduced levels of protein carbonyls by 56% and lipid peroxides by 40%, as compared with controls, but enhanced levels of low and high molecular mass thiols by 71% and 36%, respectively. The activities of superoxide dismutase, glutathione peroxidase and glutathione-S-transferase were increased in gills of goldfish exposed to 71.4mgL(-1) Sencor. At any concentration tested, Sencor did not affect the activities of glutathione reductase, glucose-6-phosphate dehydrogenase, lactate dehydrogenase or acetylcholine esterase in gills. The results of this study indicate that acute exposure of goldfish to Sencor had effect on free radical processes in gills and glutathione-dependent antioxidants effectively protect proteins and lipids from oxidation.

Toxicity of environmental Gesagard to goldfish may be connected with induction of low intensity oxidative stress in concentration- and tissue-related manners.

Prometryn is a selective herbicide commonly used in agriculture as the commercial preparation, Gesagard. Goldfish (Carassius auratus) exposure for 96h to 0.2, 1, or 5mgL(-1) Gesagard 500FW (corresponding to 0.1, 0.5, and 2.5mgL(-1) of prometryn) on indices of oxidative stress (lipid peroxides, protein carbonyls, and thiol content) and activities of antioxidant and related enzymes in gills, liver, and kidney was studied. Gills appeared to be the most resistant to Gesagard treatment, reacting to only the highest concentration of herbicide with enhanced levels of low molecular mass thiols and activities of glutathione S-transferase (GST) and glutathione reductase. Goldfish exposure to 0.2-5mgL(-1) Gesagard resulted in enhancement of carbonyl protein level and activity of superoxide dismutase (SOD), but reduced the lipid peroxide (LOOH) content and activity of glutathione peroxidase in liver. Kidney appeared to be the main target organ of Gesagard toxicity, showing the greatest number of parameters affected even under low concentrations of herbicide. An increase in the content of L-SH and activity of SOD was accompanied with decreased activities of catalase, GST, and glucose-6-phosphate dehydrogenase and reduced levels of LOOH in kidney of Gesagard treated fish. The treatment also induced various histological changes in goldfish liver and kidney which could be related to their dysfunction. The present study indicates that Gesagard induced oxidative stress of differing intensities in the three goldfish tissues and demonstrated that kidney would be the best target organ to analyze, reveal, and monitor Gesagard effects on fish.

Hepatotoxicity of herbicide Sencor in goldfish may result from induction of mild oxidative stress.

The effects of 96 h exposure to 7.14, 35.7, or 71.4 mg L(-1) of Sencor were studied on liver and plasma parameters in goldfish, Carassius auratus L. Goldfish exposure to 71.4 mg L(-1) of Sencor for 96 h resulted in a decrease in glucose concentrations in plasma and liver by 55%, but did not affect liver glycogen levels. An increase in the activity of aspartate aminotransferase, alanine aminotransferase and lactate dehydrogenase (by 24-27%, 32-72%, and 87-102%, respectively) occurred in plasma of Sencor exposed goldfish, whereas in liver activities of these enzymes decreased (by 15-17%, 19%, and 20%, respectively). Lactate concentration in plasma increased by 22-36% in all treated fish groups, whereas in liver it increased by 64% only after exposure to 35.7 mg L(-1) of Sencor. Herbicide exposure enhanced lipid peroxide levels by 49-75% and decreased activities of catalase by 46%, glutathione reductase by 25-48% and glutathione peroxidase by 21-26% suggesting development of oxidative stress in liver. The treatment induced various histological changes in goldfish liver, such as dilated sinusoids, hypertrophy and dystrophy of hepatic cells and detachment of endothelial cytoplasm with diffuse hemorrhage. The data collectively let us propose that mild oxidative stress might be responsible for the hepatotoxicity of Sencor.

Histopathological and biochemical changes in goldfish kidney due to exposure to the herbicide Sencor may be related to induction of oxidative stress.

Molecular mechanisms of toxicity by the metribuzin-containing herbicide Sencor to living organisms, particularly fish, have not yet been extensively investigated. In the present work, we studied the effects of 96 h exposure to 7.14, 35.7, or 71.4 mg L(-1) of Sencor (corresponding to 5, 25, or 50 mg L(-1) of its herbicidal component metribuzin) on goldfish (Carassius auratus L.), examining the histology, levels of oxidative stress markers, and activities of antioxidant and related enzymes in kidney as well as hematological parameters and leukocyte profiles in blood. The treatment induced various histopathological changes in goldfish kidney, such as hypertrophy of intertubular hematopoietic tissue, small and multiple hemorrhages, glomerular shrinkage, a decrease in space between glomerulus and Bowman's capsule, degeneration and necrosis of the tubular epithelium. Sencor exposure also decreased activities of selected enzymes in kidney; activities of catalase decreased by 31-34%, glutathione peroxidase by 14-33%, glutathione reductase by 17-25%, and acetylcholinesterase by 31%. However, glucose-6-phosphate dehydrogenase and lactate dehydrogenase activities increased by 25-30% and 22% in kidney after treatment with 7.14 or 35.7 mg L(-1) and 71.4 mg L(-1) Sencor, respectively. Kidney levels of protein carbonyls increased by 177% after exposure to 35.7 mg L(-1) of Sencor indicating extensive damage to proteins. Lipid peroxide concentrations also increased by 25% after exposure to 7.14 mg L(-1) of Sencor, but levels were reduced by 42% in the 71.4 mg L(-1) exposure group. The data indicate that induction of oxidative stress is one of the mechanisms responsible for Sencor toxicity to fish.