Short-term exposure to radiofrequency radiation and metabolic enzymes' activities during pregnancy and prenatal development.

Radiofrequency radiation (RFR) as an environmental and physical pollutant may induce vulnerability to toxicity and disturb fetal development. Therefore, the potential health effects of short-term mobile phone like RFR exposure (GSM 1800 MHz; 14 V/m, 2 mW/kg specific absorption rate (SAR) during 15 min/day for a week) during pregnancy and also the development of fetuses were investigated. Hepatic glucose regulation and glutathione-dependent enzymes' capacities were biochemically analyzed in adult (female) and pregnant New Zealand White rabbits. Pregnant rabbits' two-day-old offspring were included to understand their developmental stages under short-term maternal RFR exposure. We analyzed two regulatory enzymes in the oxidative phase of phosphogluconate pathways to interpret the cytosolic NADPH's biosynthesis for maintaining mitochondrial energy metabolism. Moreover, the efficiencies of maternal glutathione-dependent enzymes on both the removal of metabolic disturbances during pregnancy and fetus development were examined. Whole-body RFR exposures were applied to pregnant animals from the 15th to the 22nd day of their gestations, i.e., the maturation periods of tissues and organs for rabbit fetuses. There were significant differences in hepatic glucose regulation and GSH-dependent enzymes' capacities with pregnancy and short-term RFR exposure. Consequently, we observed that intrauterine exposure to RFR might lead to cellular ROS- dependent disturbances in metabolic activity and any deficiency in the intracellular antioxidant (ROS-scavenging) system. This study might be a novel insight into further studies on the possible effects of short-term RF exposure and prenatal development.

Protective Effects of Antioxidant Chlorophyllin in Chemically Induced Breast Cancer Model In vivo.

Glutathione-related enzymes belong to the protection mechanism of the cells against harmful oxidative damage and chemicals. Glutathione S-transferase (GST) is frequently over-expressed in various cancer cells and is involved in drug resistance. Chlorophyllin is an antioxidant molecule interfering with the GST P1-1 activity. The purpose of this study is to evaluate the short- and long-term protective effects of chlorophyllin as an antioxidant molecule on DNA damage, antioxidant enzyme activities, trace elements, and minerals in chemically induced breast cancer model in vivo. In our study, N-methyl-N-nitrosourea (MNU) was used for inducing breast carcinogenesis in female Sprague-Dawley rats. A total of 36 rats were divided into groups as short term and long term. Each group was divided into four sub-groups as control group received physiological saline solution (n = 3), Chl group (n = 5) received chlorophyllin, MNU group (n = 5) was administered MNU, and Chl + MNU group (n = 5) was treated with both chlorophyllin and MNU. Results illustrated that chlorophyllin had a significant anti-genotoxic effect in the short term, and glutathione-related enzyme activities were protected by chlorophyllin treatment in MNU-induced breast cancer model. Additionally, MNU administration impaired mineral and trace element levels including Na, Mg, K, Fe, Zn, and Co in the liver, kidney, spleen, heart, and tumor tissues; however, adverse effects of MNU were recovered upon chlorophyllin treatment in the indicated tissues of the rats. In conclusion, chlorophyllin can be used as an antioxidant molecule to ameliorate adverse effects of MNU by enhancing antioxidant enzyme activities and regulating trace element and mineral balance in several organs and tumor tissue in the breast cancer model.

Effects of donepezil on liver and kidney functions for the treatment of Alzheimer's disease.

Aim of the present study is to investigate the effects of medication with donepezil (acetylcholinesterase inhibitor) on the liver and kidney function in Alzheimer's disease (AD) and to compare the effects of donepezil medication during short (one month) and long term (six years) follow-ups. We evaluated female and male patients from Cukurova [42 AD patients; short term (5 mg/day)] and Dokuz Eylul [68 AD patients; long term (10 mg/day)] University Hospital. The results compared with the geriatric population without dementia in other words who are not in medication with donepezil. For short term evaluation all subjects underwent periodic examination with tests regarding hepatic and renal functions; firstly, before starting treatment and then repeated one month later. For long term evaluation all subjects underwent periodic examination with tests regarding hepatic and renal functions; three times at the end of each two consecutive years of treatment with donepezil. AD patients' results were also compared with 79 neurologically healthy geriatric patients without dementia who were over 65 years of age and were not receiving medication with donepezil. For this task, serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels are used to predict possible liver damage, while the blood urea nitrogen (BUN) and creatinine (CRE) levels for kidney damage. No significant difference between the groups regarding the routine control of biochemical parameters was observed in short term drug medication. In long term patients' group; the effects of two years use of donepezil on renal and hepatic function were also evaluated and levels of AST, ALT, BUN and CRE were found to be increased significantly compared to pretreatment levels. But, they remained in the reference intervals. However, levels of AST and ALT at the end of the fourth year of therapy were similar to those measured at the end of the second year, levels of BUN and CRE continuing to increase with staying below the reference limits. Functional markers obtained at the end of the sixth year of therapy were not differing from those of the fourth year. No significant difference was found during comparisons within the results of the neurologically healthy geriatric patient group. During comparisons between the two groups, measurements obtained at all-time points were significantly high in donepezil treated AD patients. We concluded that customized dosage according to hepatic and renal functions is necessary for using acetylcholinesterase inhibitor in AD patients.

Glucose-6-phosphate dehydrogenase deficiency and Alzheimer's disease: Partners in crime? The hypothesis.

Alzheimer's disease is a multifaceted brain disorder which involves various coupled irreversible, progressive biochemical reactions that significantly reduce quality of life as well as the actual life expectancy. Aging, genetic predispositions, head trauma, diabetes, cardiovascular disease, deficiencies in insulin signaling, dysfunction of mitochondria-associated membranes, cerebrovascular changes, high cholesterol level, increased oxidative stress and free radical formation, DNA damage, disturbed energy metabolism, and synaptic dysfunction, high blood pressure, obesity, dietary habits, exercise, social engagement, and mental stress are noted among the risk factors of this disease. In this hypothesis review I would like to draw the attention on glucose-6-phosphate dehydrogenase deficiency and its relationship with Alzheimer's disease. This enzymopathy is the most common human congenital defect of metabolism and defined by decrease in NADPH+H(+) and reduced form of glutathione concentration and that might in turn, amplify oxidative stress due to essentiality of the enzyme. This most common enzymopathy may manifest itself in severe forms, however most of the individuals with this deficiency are not essentially symptomatic. To understand the sporadic Alzheimer's disease, the writer of this paper thinks that, looking into a crystal ball might not yield much of a benefit but glucose-6-phosphate dehydrogenase deficiency could effortlessly give some clues.

Long-term treatment with a beta-blocker timolol attenuates renal-damage in diabetic rats via enhancing kidney antioxidant-defense system.

The factors with increasing diabetes-prevalence lead to significant global increases in chronic kidney disease. Since hyperglycemia generates more ROS and attenuates cellular antioxidant-defense mechanisms, numerous studies demonstrated that hyperglycemia-induced oxidative stress played a major role in the extracellular matrix expansion in tissues. Although no direct relation between activation of beta-adrenergic (ß-AR) system and kidney disease in diabetes and since ß-blockers demonstrate marked beneficial effects due to their scavenging free radicals and/or acting as an antioxidant in diabetic animal studies, the eventual objective of the present study was to determine whether timolol-treatment of streptozotocin-induced diabetic rats (5 mg/kg, daily following diabetes-induction, for 12-week) has advantage to prevent hyperglycemia-induced renal-damage via enhancing the depressed antioxidant defense in the kidney. Light microscopy data and their quantification demonstrated that timolol-treatment prevented basically glomerular hypertrophy, expansion in mesangium cell size, thickening and fibrosis in glomerular basement membrane, and accumulation of glycogen into tubular epithelial cells. Additionally, electron microscopy data demonstrated that timolol-treatment could also prevent diabetes-induced changes in the kidney tissue such as hypertrophy in podocytes, lost of filtration gaps and slit-diaphragms, and vacuolization in the distal tubular cells. Biochemical analysis basically on enzymes of antioxidant-defense system, including glutathione-S-transferase, glutathione reductase, and glucose-6-phosphate dehydrogenase, further supported that diabetes-induced damage in the kidney is mostly dependent on the increased oxidative stress and timolol, having an antioxidant-like action, could protect the kidney against hyperglycemia-induced damage without normalization of high-blood glucose level. Consequently, it can be suggested that although ß-blockers are widely used for the treatment of cardiovascular diseases, ß-blocker therapy of diabetics seems to be a new therapeutic approach against hyperglycemia-induced kidney damage in diabetic patients.

In vitro effects of rosmarinic acid on glutathione reductase and glucose 6-phosphate dehydrogenase.

CONTEXT: Glutathione reductase (GR, NADPH:oxidized glutathione oxidoreductase, E.C 1.6.4.2) is a flavoprotein that catalyzes the NADPH-dependent reduction of oxidized glutathione (GSSG). GR is a crucial enzyme in the antioxidant system by maintaining reduced glutathione (GSH). Glucose 6-phosphate dehydrogenase (G6PD, glucose 6-phosphate (G6P):NADP(+) oxidoreductase, EC 1.1.1.49) is the key regulatory enzyme of the pentose phosphate pathway and maintains NADPH for reductive reactions. OBJECTIVE: Rosmarinic acid (RA; α-O-caffeoyl-3,4-dihydroxyphenyl lactic acid) is an ester of caffeic acid (CA) and 3,4-dihydroxyphenyllactic acid. It has a number of interesting biological activities. The inhibiting activities of the RA on GR and G6PD are investigated here for the first time. MATERIALS AND METHODS: GR and G6PD were purified from tissues, then the effects of RA are investigated. RESULTS: This study reports that RA, which was isolated from Echium vulgare L. (Boraginaceae), inhibits purified GR and G6PD in a concentration-dependent manner. Kinetic characterizations and inhibition constants are investigated. DISCUSSION AND CONCLUSION: Because of their importance in the antioxidative defense system, investigation of the inhibitors of these enzymes is important for drug development.

Comparative in vitro effects of some metal ions on bovine kidney cortex glutathione reductase.

Heavy metal pollution can arise from many sources and damage many organisms. Exposure to the metal ions can leads to a reduction in cellular antioxidant enzyme activities and lowers cellular defense against oxidative stress. In this study we have tested effects of the some metal ions on the purified bovine kidney cortex glutathione reductase (GR). Cadmium (Cd²+), nickel (Ni²+), and zinc (Zn²+) showed inhibitory effect on the enzyme. The obtained IC50 values of Cd²+, Ni²+, and Zn²+ are 0.027, 0.8, and 1 mM, respectively. Kinetic characterization of the inhibition is also investigated. Cd²+ inhibition is noncompetitive with respect to both oxidized glutathione (GSSG) (Ki(GSSG) 0.060 ± 0.005 mM) and NADPH (Ki(NADPH) 0.025 ± 0.002 mM). Ni²+ inhibition is noncompetitive with respect to GSSG (Ki(GSSG) 0.329 ± 0.016 mM) and uncompetitive with respect to NADPH (Ki(NADPH) 0.712 ± 0.047 mM). The effect of Zn²+ on GR activity is consistent with noncompetitive inhibition pattern when the varied substrate is the GSSG (Ki(GSSG) 0.091 ± 0.005 mM) and the NADPH (Ki(NADPH) 0.226 ± 0.01 mM), respectively. GR inhibition studies may be useful for understanding the mechanisms for oxidative damage associated with heavy metal toxicity.

A comparative study with colchicine on glutathione reductase.

Colchicine is a drug used for the treatment of FMF, primary biliary cirrhosis, psoriasis, Behçet's disease, aphthous stomatitis. Glutathione reductase (GR; E.C 1.6.4.2) is a crucial enzyme which reduces glutathione disulphide to the sulfhydryl form GSH by the NADPH-dependent reduction, which is an important cellular antioxidant system. The purpose of the present work is to evaluate the in vitro effects of colchicine on GR from various sources. The component of glutathione redox cycle, GR, plays important role in the protection of the cell from the toxic effects of reactive oxygen species. Due to its significance the enzyme has been purified from a number of animals, plants and microbial sources and studied the in vitro effects of many chemical compounds or drugs on enzyme activity. We have established that colchicine inhibits GR in a concentration dependent manner. We have investigated the kinetic characterization, inhibition types and constants (Ki).

Inhibition of purified bovine liver glutathione reductase with some metal ions.

Glutathione reductase (GR; E.C. 1.6.4.2) is a flavoprotein that catalyzes the NADPH-dependent reduction of oxidized glutathione (GSSG). In this study we tested the effects of Al3+, Ba2+, Ca2+, Li+, Mn2+, Mo6+, Cd2+, Ni2+, and Zn2+ on purified bovine liver GR. In a range of 10 microM-10 mM concentrations, Al3+, Ba2+, Li+, Mn2+, and Mo6+, and Ca2+ at 5 microM-1.25 mM, had no effect on bovine liver GR. Cadmium (Cd2+), nickel (Ni2+), and zinc (Zn2+) showed inhibitory effects on this enzyme. The obtained IC50 values of Cd2+, Ni2+, and Zn2+ were 0.08, 0.8, and 1 mM, respectively. Cd2+ inhibition was non-competitive with respect to both GSSG (Ki(GSSG) 0.221 +/- 0.02 mM) and NADPH (Ki(NADPH) 0.113 +/- 0.008 mM). Ni2+ inhibition was non-competitive with respect to GSSG (Ki(GSSG) 0.313 +/- 0.01 mM) and uncompetitive with respect to NADPH (Ki(NADPH) 0.932 +/- 0.03 mM). The effect of Zn2+ on GR activity was consistent with a non-competitive inhibition pattern when the varied substrates were GSSG (Ki(GSSG) 0.320 +/- 0.018 mM) and NADPH (Ki(NADPH) 0.761 +/- 0.04 mM), respectively.

The inhibition kinetics of yeast glutathione reductase by some metal ions.

Glutathione reductase (GR, type IV, Baker's yeast, E.C 1.6.4.2) is a flavoprotein that catalyzes the NADPH-dependent reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH). In this study some metal ions have been tested on GR; lithium, manganese, molybdate, aluminium, barium, zinc, calcium, cadmium and nickel. Cadmium, nickel and calcium showed a good to moderate inhibitory effect on yeast GR. GR is inhibited non-competitively by Zn2+ (up to 2 mM) and activated above this concentration. Ca2+ inhibition was non-competitive with respect to GSSG and uncompetitive with respect to NADPH. Nickel inhibition was competitive with respect to GSSG and uncompetitive with respect to NADPH. The inhibition constants for these metals on GR were determined. The chelating agent EDTA recovered 90% of the GR activity inhibited by these metals.

Gender related differential effects of Omega-3E treatment on diabetes-induced left ventricular dysfunction.

The present study was designed to determine whether there are beneficial effects of intake of Omega-3E (containing 70% pure omega-3 and 2% natural vitamin E) in cardiac dysfunction of diabetic rats. We also examined whether there are gender-related differences in the responses to the intake of Omega-3E on the heart dysfunction. Experiments were performed by using Langendorff-perfused hearts from normal, diabetic (with 50 mg/kg streptozotocin), and Omega-3E (50 mg/kg body weight/day) treated diabetic 3-month-old Wistar rats. Omega-3E treatment of the diabetics caused small, but significant decrease (13% and 14% female versus male) in the blood glucose level. Omega-3E treatment of the diabetic female rats did not prevent diabetes-induced decrease in left ventricular developed pressure (LVDP) and increase in left ventricular end-diastolic pressure (LVEDP) with respect to the control female rats. On the other hand, the treatment of diabetic male rats caused significant recovery in depressed LVDP. Furthermore, such treatment of diabetic female and male rats caused significant recovery in depressed rates of changes of developed pressure. This effect was more significant in males. Besides, Omega-3E caused significant further lengthening in the diabetes-induced increased time to the peak of the developed pressure in females, while it normalized the lengthening in the relaxation of the developed pressure in diabetic males. In addition, Omega-3E treatment caused significant restorations in the diabetes-induced altered activities of antioxidant enzymes without any significant gender discrepancy. Present data show that there are gender related differences in diabetic heart dysfunction and the response to antioxidant treatment.

Purification and kinetic properties of glutathione reductase from bovine liver.

Glutathione reductase (GR, NADPH: oxidized glutathione oxidoreductase, EC 1.6.4.2) catalyzes the reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH) using NADPH as reducing cofactor. The aim of the present work was to purify and characterize GR from bovine liver. GR was purified using 2', 5' ADP-Sepharose 4B and DEAE-Sepharose Fast Flow columns. The enzyme has been purified 5456-fold and with a yield of 38.4%. The molecular and catalytic properties of bovine liver GR have been studied. Optimum temperature and pH was found to be 50 degrees C and 7, respectively. The activation energy of the reaction catalyzed by the enzyme was 9.065 kcal/mole. The molecular weight of the enzyme was found to be 55 kDa by SDS-PAGE. Kinetic characterization of bovine liver GR was also investigated, Km(NADPH) 0.063 +/- 0.008 mM and Km(GSSG) 0.154 +/- 0.015 mM were determined. It is accepted that parallel lines observed in these double reciprocal plots obeys Ping Pong mechanism and we have showed this in our steady state study. According to our results of statistical analysis, the Ping Pong mechanism is a suitable model since the loss function is less than the other mechanisms. However, competitive inhibition by a product could be accepted in sequential mechanisms but not in a Ping Pong mechanism. In this study, kinetic data are consistent with a branching reaction mechanism previously proposed for GR from other sources by other studies.

Selenium treatment protects diabetes-induced biochemical and ultrastructural alterations in liver tissue.

We have shown that a single dose of streptozotocin (STZ) (50 mg/kg body weight) injected into rats caused significant changes in some antioxidant enzyme activities, such as glutathione peroxidase, glutathione reductase, glutathione-S-transferase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase activities, and acid-soluble sulfhydryl levels of the liver tissue with respect to the control rats. Furthermore, these alterations in the activities of the antioxidant enzymes were accompanied by significant changes in the ultrastructure of the liver tissue; mainly intercellular biliary canaliculi were distended and contained stagnant bile, swollen mitochondria in hepatocytes and disoriented and disintegrating cristae, dilatation of the rough endoplasmic reticulum (rER) with detachment of ribosomes, and dissociation of polysomes. Both diabetic and normal rats were treated with sodium selenite (5 micromol/kg/d, intra peritoneally) for 4 wk following 1 wk of diabetes induction. This treatment of diabetic rats improved significantly diabetes-induced alterations in liver antioxidant enzymes. Moreover, treating of diabetic rats with sodium selenite prevented primarily the variation in staining quality of hepatocytes nuclei, increased density and eosinophilia of the cytoplasm, focal sinusoidal dilatation and congestion, and increased numbers of mitochondria with different size and shape. In summary, treatment of diabetic rats with sodium selenite has beneficial effects on both antioxidant system and the ultrastructure of the liver tissue. These findings suggest that diabetes-induced oxidative stress can be responsible for the development of diabetic complications and antioxidant treatment can protect the target organs against diabetes.

Total calcium content of sacs associated with inguinal hernia, hydrocele or undescended testis reflects differences dictated by programmed cell death.

INTRODUCTION: Inguinal hernia and hydrocele are suggested to result from the persistence of smooth muscle (SM) which should undergo programmed cell death (PCD) after presenting transiently to propel the testis. Since Ca(2+) is involved in PCD, the Ca(2+) contents of the peritoneum and sacs associated with undescended testis, inguinal hernia and hydrocele were determined and compared. MATERIALS AND METHODS: Sacs were obtained from boys with undescended testis (n = 11), inguinal hernia (n = 22) and hydrocele (n = 10), and girls with inguinal hernia (n = 7). The calcium content of the sacs and peritoneal samples (n = 6) was determined through atomic absorption spectrophotometry. Calcium contents were compared according to their sources using the Mann-Whitney U test and p values of <0.05 were considered significant. RESULTS: While revealing similar Ca(2+) contents as the peritoneum, sacs associated with undescended testis and hydrocele contained more Ca(2+) contents than the sacs of boys and girls with inguinal hernia (p < 0.05). CONCLUSIONS: Sacs associated with inguinal hernia, which are known to contain SM all around the mesothelial layer, contain the least Ca(2+). Despite the decrease in SM, sacs associated with hydrocele contain more Ca(2+). Since PCD is associated with Ca(2+) overload and inhibition of Ca(2+) load inhibits PCD, differences in Ca(2+) content may reflect the inhibition of PCD at different stages and for different reasons in inguinal hernia or hydrocele of childhood.

Less calcium in cremaster muscles of boys with undescended testis supports a deficiency in sympathetic innervation.

In addition to an increase in contractility, contracted fibers and small and electron-dense mitochondria have suggested an increase of cytosolic Ca(2+) within the cremaster muscles (CM) associated with undescended testis. Therefore, the Ca(2+) content of CM associated with an undescended testis was determined and compared with the Ca(2+) content associated with inguinal hernia and hydrocele and in internal oblique muscles. CM samples from boys with undescended testis (n = 9), inguinal hernia (n = 15), and hydrocele (n = 7) and from girls with inguinal hernia (n = 8) were obtained. The calcium contents of these samples and of samples from internal oblique muscle (n = 6) were determined by atomic absorption spectrophotometry. While the Ca(2+) contents of CM from boys with undescended testis and from girls with inguinal hernia were similar (p > 0.05), the internal oblique muscle and the CM from boys with inguinal hernia and hydrocele contained more Ca(2+) than CM from boys with undescended testis (p < 0.05). Despite evidence of an increase in cytosolic calcium, the decrease in total calcium content suggests a decrease in the adenylyl cyclase activity, thus inhibition of influx of Ca(2+), but an increase in phospholipase C activity, and generation of more inositol 1,4,5-trisphosphate and mobilization of calcium from internal stores. Since the sympathetic system acts through adenylyl cyclase in striated muscles and is sexually dimorphic, similar Ca(2+) contents as encountered in girls suggest an alteration in the balance of autonomic innervation against the sympathetic system in boys with undescended testis.

Abietic acid inhibits lipoxygenase activity.

In this study, it was shown that abietic acid, an abietane diterpenoid, inhibited soybean 5-lipoxygenase (linoleate: oxygen oxidoreductase, EC 1.13.11.12) and an IC(50) of 29.5 +/- 1.29 microM was determined. Since the lipoxygenase pathway leads to the biosynthesis of leukotrienes this result supports the view that abietic acid may be used in the treatment of allergic reactions.