Hepatic GSH Content Research Articles

English

The present study tries to investigate oxidative stress - hyperglycaemia relationship through studying role of ROS and oxidative stress in the pathogenesis of type 2 diabetes mellitus and role of NAC as antioxidant in normalizing that disorders. 80 rats were randomly assigned into four equal groups, 20 rats each; Control group (C) fed on the balanced ration and ordinary drinking water ad-libitum, Fructose fed group (F) fed on the balanced ration and fructose 10% (100g/L) in drinking water, Fructose and N-acetylcysteine group (FN) fed on the balanced ration and fructose 10% (100g/L) in drinking water for 4 weeks then injected I/P with N-acetylcysteine in dose of 200 mg/Kg b.wt. daily for the last six weeks and N-acetylcysteine and Fructose group (NF) fed on the balanced ration and ordinary drinking water and injected I/P with N-acetylcysteine for 4 weeks then given fructose in drinking water for the last six weeks. Blood samples and liver specimens were collected from ten animals from each group after 2 and 10 weeks. The obtained results revealed that fructose administration increased hepatic MDA content and GSPx activity and plasma insulin, insulin resistance and glucose levels. Conversely, excess fructose decreased hepatic GSH content, expression of insulin-receptor's protein gene, SOD, hexokinase and G6PD activities. Those disorders originated a case of type 2 diabetes. Injection of NAC either as treatment or protective relieved all that disorders towards normal condition through its antioxidant influ- ence.

Role of connexin 32 in acetaminophen toxicity in a knockout mice model

Gap junctional intercellular communication (GJIC), by which glutathione (GSH) and inorganic ions are transmitted to neighboring cells, is recognized as being largely involved in toxic processes of chemicals. We examined acetaminophen (APAP)-induced hepatotoxicity clinicopathologically using male wild-type mice and mice lacking the gene for connexin32, a major gap junction protein in the liver [knockout (Cx32KO) mice]. When APAP was intraperitoneally administered at doses of 100, 200, or 300mg/kg, hepatic centrilobular necrosis with elevated plasma aminotransferase activities was observed in wild-type mice receiving 300mg/kg, and in Cx32KO mice given 100mg/kg or more. At 200mg/kg or more, hepatic GSH and GSSG contents decreased significantly and the effect was more severe in wild-type mice than in Cx32KO mice. On the other hand, markedly decreased GSH staining was observed in the hepatic centrilobular zones of Cx32KO mice compared to that of wild-type mice. These results demonstrate that Cx32KO mice are more susceptible to APAP hepatotoxicity than wild-type mice, and indicate that the distribution of GSH of the centrilobular zones in the hepatic lobules, rather than GSH and GSSG contents in the liver, is important in APAP hepatotoxicity. In conclusion, Cx32 protects against APAP-induced hepatic centrilobular necrosis in mice, which may be through the GSH transmission to neighboring hepatocytes by GJIC.

Nilotinib Interferes with the Signalling Pathways Implicated in Acetaminophen Hepatotoxicity

Nilotinib, a second-generation tyrosine kinase inhibitor, has been recently approved for the treatment for chronic myeloid leukaemia. The objective of this study was to explore the potential effects of clinically relevant doses of nilotinib against acetaminophen (APAP)-induced hepatotoxicity in mice. To simulate the clinical application in human beings, nilotinib (25 and 50 mg/kg) was administered to mice 2 hr after APAP intoxication (500 mg/kg). The results indicated that nilotinib (25 mg/kg) (i) abolished APAP-induced liver injury and necro-inflammation, (ii) increased hepatic-reduced glutathione (GSH) and its related enzymes synthesis, (iii) suppressed hepatic oxidative/nitrosative stress cascades, (iv) decreased neutrophil accumulation in the liver, and (v) prevented the over-expression of B-cell lymphoma-2 (bcl-2), cyclin-D1 and stem cell factor receptor (c-Kit) proteins in the liver. Although nilotinib (50 mg/kg) acted through the same mechanisms, there was severe depletion in hepatic GSH content by nilotinib itself at that dose level, rather than the potent stimulation observed by using a dose of 25 mg/kg. Consequently, the mortality rate after 18 hr was 100% for nilotinib (50 mg/kg) + APAP (750 mg/kg) versus 60% for APAP (750 mg/kg) and 40% for nilotinib (25 mg/kg) + APAP (750 mg/kg) in the survival analysis experiment. In conclusion, nilotinib can counteract the hepatotoxicity produced by a non-lethal dose of APAP. However, there is a risk of aggravating the mortality for a lethal dose of APAP when nilotinib is co-administered at doses relatively high, or near to the clinical range because of hepatic GSH depletion and c-kit inhibition.

Antioxidant capacity responsible for a hypocholesterolemia is independent of dietary cholesterol in adult rats fed rice protein

Dietary cholesterol and aging are major risk factors to accelerate oxidation process for developing hypercholesterolemia. The major aim of this study is to elucidate the effects of rice protein on cholesterol level and oxidative stress in adult rats fed with and without cholesterol. After 2weeks of feeding, hepatic and plasma contents of cholesterol, reduced glutathione (GSH), oxidized glutathione (GSSG), malondialdehyde (MDA) and protein carbonyl (PCO) were measured. In liver, total antioxidative capacity (T-AOC), activities of antioxidant enzymes (total superoxide dismutase, T-SOD; catalase, CAT), glutathione metabolizing enzyme activities and gene expression levels (γ-glutamylcysteine synthetase, γ-GCS; glutathione reductase, GR; glutathione peroxidase, GPx) were determined. Under cholesterol-free/enriched dietary condition, T-AOC, activities of T-SOD and CAT, glutathione metabolism related enzymes' activities and mRNA levels (γ-GCS, GR and GPx) were effectively stimulated by rice proteins as compared to caseins. Compared with caseins, rice proteins significantly increased hepatic and plasma GSH contents, whereas hepatic and plasma accumulations of MDA, PCO and GSSG were significantly reduced by rice protein-feedings. As a result, the marked reductions of cholesterol in the plasma and in the liver were observed in adult rats fed rice proteins with and without cholesterol. The present study demonstrates that the hypocholesterolemic effect of rice protein is attributable to inducing antioxidative response and depressing oxidative damage in adult rats fed cholesterol-free/enriched diets. Results suggest that the antioxidant capability involved in the hypocholesterolemic action exerted by rice protein is independent of dietary cholesterol during adult period.

Effects of dietary concentrations of methionine on growth performance and oxidative status of broiler chickens with different hatching weight

1. A study was conducted to evaluate the effects of two hatching weight (HW) levels and two dietary concentrations of methionine on the growth performance and oxidative status of broilers. Male Arbor Acres chickens were divided into two groups on their HW (low and high HW, H and L). Each HW group was then distributed into two subgroups, of similar HW, receiving either low or high dietary concentrations of methionine (4.9 g methionine/kg, LM; 5.9 g methionine/kg, HM). Thus, all day-old birds were distributed into 4 treatments (H-LM, H-HM, L-LM, L-HM) × 6 replicates × 10 birds for 21 d. 2. Broilers with high HW were heavier than those with low HW during the 21 d assay, which appeared to result from increased body weight gain rather than improved feed conversion efficiency. A higher dietary concentration of methionine (5.9 g/kg) improved growth performance of broilers with low HW in terms of body weight gain and feed conversion ratio. 3. Broilers with different HW had similar antioxidant status both in serum and liver. 4. Broilers given a diet containing 5.9 g/kg methionine had enhanced serum superoxide dismutase (SOD) activity and decreased hepatic malondialdehyde (MDA) content at day 7. 5. Broilers given a diet containing 5.9 g/kg methionine had a higher hepatic reduced glutathione (GSH):glutathione disulphide (GSSG) ratio than those given a diet containing 4.9 g/kg methionine at day 21. High dietary methionine concentration reduced hepatic GSH content and glutathione peroxidase (GPX) activity of broilers with high HW at day 7 and at day 21, respectively, but increased hepatic GSH content of broilers with low HW at day 7. 6. Although broilers with different HW had similar oxidative status as indicated by several parameters in blood and liver, HW can have positive effects on the subsequent growth performance of broilers, and a higher dietary methionine concentration (5.9 g/kg) can improve growth performance and antioxidant status in broilers exhibiting low HW.

Glutathione defense mechanism in liver injury: Insights from animal models

Glutathione (GSH) is the most abundant cellular thiol antioxidant and it exhibits numerous and versatile functions. Disturbances in GSH homeostasis have been associated with liver diseases induced by drugs, alcohol, diet and environmental pollutants. Until recently, our laboratories and others have developed mouse models with genetic deficiencies in glutamate-cysteine ligase (GCL), the rate-limiting enzyme in the GSH biosynthetic pathway. This review focuses on regulation of GSH homeostasis and, specifically, recent studies that have utilized such GSH-deficient mouse models to investigate the role of GSH in liver disease processes. These studies have revealed a differential hepatic response to distinct profiles of hepatic cellular GSH concentration. In particular, mice engineered to not express the catalytic subunit of GCL in hepatocytes [Gclc(h/h) mice] experience almostcomplete loss of hepatic GSH (to 5% of normal) and develop spontaneous liver pathologies characteristic of various clinical stages of liver injury. In contrast, mice globally engineered to not express the modifier subunit of GCL [Gclm⁻/⁻ mice] show a less severe hepatic GSH deficit (to ≈15% of normal) and exhibit overall protection against liver injuries induced by a variety of hepatic insults. Collectively, these transgenic mouse models provide interesting new insights regarding pathophysiological functions of GSH in the liver.

Significance of alterations in the metabolomics of sulfur-containing amino acids during liver regeneration

It has been known that liver regeneration is accompanied with a profound change in the metabolomics of sulfur-containing substances in liver. However, its physiological significance in the liver regenerative process is still unclear. Our previous work showed that buthioninesulfoximine and phorone, both widely used to deplete intracellular glutathione (GSH) in biological experiments, induced contrasting changes in the sulfur-containing amino acid metabolism in liver. In this study we employed these GSH-depleting agents to evaluate the role of sulfur-containing substances in the early phase of liver regeneration. Male rats treated with buthioninesulfoximine or phorone were subjected to two-thirds partial hepatectomy (PHx). At the doses used, the magnitude of GSH depletion after PHx was comparable, but buthioninesulfoximine administration inhibited the progression of liver regeneration as determined by liver weight increase, elevation of serum alanine aminotransferase activity, and cyclin D1 and proliferating cell nuclear antigen (PCNA) protein expressions, whereas liver recovery was significantly accelerated in the phorone-treated rats, suggesting that the role of GSH in this process is minimal. Hepatic concentrations of methionine, S-adenosylmethionine, cysteine, taurine and GSH were all elevated by PHx. Methionine adenosyltransferase activity was also induced in the remnant liver. Buthioninesulfoximine administration depressed the elevation of S-adenosylmethionine, but increased the catabolism of cysteine to taurine. In contrast, S-adenosylmethionine elevation was augmented whereas cysteine, hypotaurine and taurine were decreased in the phorone-treated rats. PHx elevated hepatic putrescine and spermidine, but lowered spermine concentrations. Buthioninesulfoximine administration increased putrescine further, but decreased spermidine and spermine concentrations. On the contrary, both spermidine and spermine concentrations were elevated in the rats treated with phorone. The results suggest that the availability of S-adenosylmethionine plays a critical role in the progression of liver regeneration via enhancement of polyamine synthesis. These findings raise the possibility that regulating hepatic transsulfuration reactions may be capable of modifying the recovery process after liver injury.

Application of Interleukin-22 Mediates Protection in Experimental Acetaminophen-Induced Acute Liver Injury

Acetaminophen (APAP, paracetamol)-induced hepatotoxicity, although treatable by timely application of N-acetylcysteine, can be fatal. Because it is among the common causes of acute liver failure in intensive care units and in light of its gradually increasing incidence, the need for novel therapeutic strategies aimed at severe intoxication is apparent. Recently, it has been shown that IL-22, a STAT3-activating cytokine, has the capability to mediate liver protection. Herein, the protective potential of IL-22 in murine APAP-induced hepatotoxicity was assessed. Intravenous administration of prophylactic IL-22 significantly reduced serum alanine aminotransferase levels and histopathologic damage in APAP-induced liver injury, a process that coincided with increased hepatocyte proliferation invivo. Concomitant gene expression analysis revealed hepatic induction of genes prototypically up-regulated by the IL-22/STAT3 axis, among others suppressor of cytokine signaling-3, lipocalin-2, and α1-antichymotrypsin. Notably, in a translational setting of therapeutic treatment 2 hours after APAP, IL-22 supported protection in the context of suboptimal N-acetylcysteine dosing. IL-22 likewise connected to augmented hepatocyte proliferation in this experimental setting. As detected by analysis of inflammatory cytokine production, systemically applied IL-22 did not display acute immunomodulation/stimulation in otherwise untreated or endotoxemic mice. Those latter observations clearly confirm acute tolerability of systemically applied IL-22. Observations presented altogether suggest that therapeutic IL-22 administration is a conceivable tissue-protective regimen aimed at hard-to-treat patients with severe APAP-induced hepatotoxicity.

The hepatoprotection of caffeic acid and rosmarinic acid, major compounds of Perilla frutescens, against t-BHP-induced oxidative liver damage

Perilla frutescens leaves are often used in East Asian gourmet food. In this study, we investigated the hepatoprotective effects of caffeic acid (CA), rosmarinic acid (RA), and their combination. P. frutescens contains 1.32μg CA/mg dry material (DM) and 26.84μg RA/mg DM analyzed by HPLC-DAD and HPLC-MS. CA remarkably reduced the oxidative damage than rosmarinic acid in an in vitro study. Oral intubation with CA or RA alone for five days was conducted prior to treatment with a single dose of tert-butyl hydroperoxide (0.5mmol/kg b.w., i.p.), which led to a significant reduction of indicators of hepatic toxicity, such as aspartate aminotransferase, alanine aminotransferase, oxidized glutathione, lipid peroxidation and enzyme activities related to antioxidant such as catalase, glutathione peroxidase and superoxide dismutase. Interestingly, compared to treatment with CA or RA alone, a combination of both compounds more increased the endogenous antioxidant enzymes and glutathione (GSH) and decreased lipid peroxidation in livers. These results suggest that CA from perilla leaves plays a role in the increased hepatic GSH concentration, and shows an additive hepatic protection with RA against oxidative hepatic damage.

Protective effect of apigenin on mouse acute liver injury induced by acetaminophen is associated with increment of hepatic glutathione reductase activity

Apigenin, a natural plant flavone, has many beneficial effects, but there is no report about treatment of acetaminophen-induced liver injury. Our aim was to examine the protective effect of apigenin on acetaminophen-induced mouse acute liver injury and to investigate the potential mechanisms. A mouse model with acute liver injury was induced by intraperitoneally given acetaminophen 350 mg kg(-1) after oral administration of apigenin 100 and 200 mg kg(-1) for 7 days. The results showed that after treatment with apigenin, the levels of serum alanine aminotransferase and aspartate aminotransferase were gradually decreased, and the severity of liver injury was decreased. In particular, significant changes in liver necrosis were observed in the apigenin 200 mg kg(-1) group. Apigenin could gradually increase the hepatic glutathione reductase (GR) activity and reduced glutathione (GSH) content, and decrease the hepatic malondialdehyde content, but the activities of glutathione peroxidase and glutathione S-transferase in hepatic tissues between the model group and the apigenin-treated groups were not significantly different. It was concluded that apigenin could protect against acetaminophen-induced acute liver injury in mice, and the mechanisms might be associated with enhancing hepatic GSH content via increment of GR activity.

Exposure to bifenthrin causes immunotoxicity and oxidative stress in male mice

Bifenthrin (BF) is one of the most commonly used pesticides among the synthetic pyrethroids. The effects of BF exposure on the induction of immunotoxicity and oxidative stress were studied both in adolescent and adult male ICR mice. Both the weights of the spleen and thymus decreased significantly in the adolescent mice when they were treated with 20 mg/kg BF for 3 weeks. We found that the 3-week oral administration of BF during puberty increased the transcriptional levels of the genes TNF and IL2 in the spleen and IL2 as well as IL4 in the thymus. The effect of BF exposure on the induction of oxidative stress was also studied in serum and liver samples. The total antioxidant capacity and activity of superoxide dismutase were altered significantly in the serum of the 20 mg/kg BF-treated adolescent mice, and the activity of glutathione peroxidase (GPX) decreased significantly in the serum of adolescent and adult mice after 3 weeks of oral administration of 20 mg/kg BF. Compared to serum, hepatic GSH content increased significantly in both the adolescent and adult mice exposed to 20 mg/kg BF; hepatic CAT and GPX activities were altered significantly, even in adolescent mice, after treatment with 10 mg/kg BF. Taken together, the results of this study suggest that exposure to BF, especially during puberty, has the potential to induce immunotoxicity accompanied by oxidative stress in male mice. These findings will help in elucidating the mechanism of toxicity induced by BF in mice.

Chimeric mice with a humanized liver as an animal model of troglitazone-induced liver injury

Troglitazone (Tro) is a thiazolidinedione antidiabetic drug that was withdrawn from the market due to its association with idiosyncratic severe liver injury. Tro has never induced liver injury in experimental animals in vivo. It was assumed that the species differences between human and experimental animals in the pharmaco- or toxicokinetics of Tro might be associated with these observations. In this study, we investigated whether a chimeric mouse with a humanized liver that we previously established, whose replacement index with human hepatocytes is up to 92% can reproduce Tro-induced liver injury. When the chimeric mice were orally administered Tro for 14 or 23 days (1000mg/kg/day), serum alanine aminotransferase (ALT) was significantly increased by 2.1- and 3.6-fold, respectively. Co-administration of l-buthionine sulfoximine (10mM in drinking water), an inhibitor of glutathione (GSH) synthesis, unexpectedly prevented the Tro-dependent increase of ALT, which suggests that the GSH scavenging pathway will not be involved in Tro-induced liver injury. To elucidate the mechanism of the onset of liver injury, hepatic GSH content, the level of oxidative stress markers and phase I and phase II drug metabolizing enzymes were determined. However, these factors were not associated with Tro-induced liver injury. An immune-mediated reaction may be associated with Tro-induced liver toxicity in vivo, because the chimeric mouse is derived from an immunodeficient SCID mouse. In conclusion, we successfully reproduced Tro-induced liver injury using chimeric mice with a humanized liver, which provides a new animal model for studying idiosyncratic drug-induced liver injury.

Protective effect of l-theanine on carbon tetrachloride-induced acute liver injury in mice

We studied effects of l-theanine, a unique amino acid in tea, on carbon tetrachloride (CCl4)-induced liver injury in mice. The mice were pre-treated orally with l-theanine (50, 100 or 200mg/kg) once daily for seven days before CCl4 (10ml/kg of 0.2% CCl4 solution in olive oil) injection. l-theanine dose-dependently suppressed the increase of serum activity of ALT and AST and bilirubin level as well as liver histopathological changes induced by CCl4 in mice. l-theanine significantly prevented CCl4-induced production of lipid peroxidation and decrease of hepatic GSH content and antioxidant enzymes activities. Our further studies demonstrated that l-theanine inhibited metabolic activation of CCl4 through down-regulating cytochrome P450 2E1 (CYP2E1). As a consequence, l-theanine inhibited oxidative stress-mediated inflammatory response which included the increase of TNF-α and IL-1β in sera, and expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in livers. CCl4-induced activation of apoptotic related proteins including caspase-3 and PARP in mouse livers was also prevented by l-theanine treatment. In summary, l-theanine protects mice against CCl4-induced acute liver injury through inhibiting metabolic activation of CCl4 and preventing CCl4-induced reduction of anti-oxidant capacity in mouse livers to relieve inflammatory response and hepatocyte apoptosis.

Evidence of Reduced Feeding and Oxidative Stress in Common Tree Frogs (Hypsiboas pulchellus) from an Agroecosystem Experiencing Severe Drought

The intensification of modern agriculture may impact amphibian populations through habitat loss and the direct and indirect effects of the pesticides upon which it relies. The increased homogeneity of modern agricultural landscapes may be detrimental in times of extreme low and high temperatures associated with climate change, as refuge abundance and habitat connectivity decrease. In this study, we evaluated the impacts of the herbicide glyphosate and subsequent intense drought on the Common Tree Frog, Hypsiboas pulchellus, inhabiting an agricultural landscape. We examined a series of organismic indices (stomach content index, hepatosomatic index, body fat index, gonadosomatic index, condition factor) as well as biomarkers of oxidative stress (hepatic catalase activity and reduced glutathione [GSH] content), exposure to contaminants (hepatic gluthatione-S-transferase activity), and genotoxicity (frequency of micronuclei). No significant differences were observed in the parameters measured when comparing frogs sampled before, 2, and 15 days after glyphosate exposure. However, anurans sampled in the same site two months later, when a drought was at its peak, presented a decrease in stomach content and hepatosomatic index, as well as an increase in hepatic catalase activity, hepatic GSH content and micronuclei frequency in peripheral circulating erythrocytes. Our findings demonstrate that drought is challenging to these anurans in this environment as evidenced by an apparent reduction in food intake and oxidative stress.

Evaluation of the effect of Spirulina against Gamma irradiation induced oxidative stress and tissue injury in rats

Many antioxidants have been investigated as hepato-protectors against ionizing radiation induced injury since they reduce the oxidative effect of the reactive oxygen species on normal cells . Spirulina is a potent scavenger of a variety of free radicals. The aim of this study was to investigate the radio-protective effect of Spirulina algae a gainst oxidative stress and tissue injury caused by gamma radiation. Rats were irradiated at two doses of 2 and 4Gy from cesium-137 source. Ten days prior to irradiation, animals received Spirulina daily (300mg/kg body weight i.p.). In the irradiated animal group, the oxidative stress markers malondialdehyde (MDA) was significantly increased in the liver, while a marked decrease in hepatic contents of DNA, and glutathione (GSH).The level of cholesterol, triglyceride (TG), low density lipoprotein (LDL), as well as activitiy of aspartate aminotransferase (AST), alkaline phosphatase (ALP), and gamma-glutamyl transferase (GGT), were significantly increased in sera of the irradiated rats. This was coupled with a decreased serum level of high-density lipoprotein (HDL) and total serum protein by irradiation. The administration of Spirulina alone daily for 10 days caused a significant decrease in MDA and produced significant elevation of liver GSH. Moreover, a significant decrease occurred in cholesterol, and TG level with no change in serum LDL or HDL levels in serum. Treatment of rats with Spirulina for 10 days before acute irradiation significantly abolished radiation induced elevation in liver MDA level and significantly maintained hepatic GSH content and CAT activity close to the control values. Pre-irradiation treatment of rats with Spirulina showed a significantly higher hepatic DNA content compared to that of irradiated rats. The level of cholesterol, TG, HDL, LDL as well as the activity of AST, ALT, and GGT in serum was significantly ameliorated when Spirulina was injected before irradiation. In conclusion, the increase in oxidative stress markers and the concomitant change in antioxidant levels indicate the role of oxidative stress in radiation-induced tissue damage. It was concluded that administration of Spirulina algae possess a radio protective capacity against ionizing-radiation induced oxidative stress and organ injury.

Systemic and Hepatic Thiol Redox Dysregulation in Graft Versus Host Disease

Graft-versus-host disease (GVHD) is mediated by a series of pathophysiological events that follow transplant conditioning and are amplified by cellular effectors and inflammatory pathways. In addition, GVHD increases oxidative stress and alters the redox environment. Whether redox dysregulation in itself potentiates the tissue injury of GVHD is unknown. We hypothesized that redox pertubation is a proximal event that precedes target organ injury in GVHD. To explore this, we applied mass-spectrometry based thiol redox metabolomics to analyze the plasma and hepatic sulfur amino acid (SAA) and thiol oxidation states in several well characterized murine models of GVHD (Allo) and compared to syngeneic (Syn) controls, at multiple time-points post-transplantation. Of the 38 amino-acid metabolites profiled, significant depletion of plasma glutathione (GSH) redox potential was noted in Allo (-127.3 ± 16 mV) compared to Syn [158.1±18 mV (p = 0.04)] mice as early as day +4 post- transplantation. This change in GSH redox state was not reflected in the cysteine (Cys/CySS) system, the principal redox couple in plasma. By day +14 post-transplantation, further decline in plasma GSH was noted in Allo mice while normalization of this metabolite was seen in Syn controls. The total hepatic GSH concentration in Allo (43.2 ± 13 nmol/mg proteins) declined by ∼33% (p = 0.05) relative to Syn group (68 ± 16 nmol/mg proteins), indicating impaired GSH synthesis. Additionally, a significant twofold (p = 0.0007) rise in hepatic homocysteine and ophthalmic acid levels were detected in Allo mice, linking both SAA and redox metabolic imbalance to GVHD. To investigate mechanism(s) of GSH redox loss in GVHD, hepatic mRNA and protein levels of gamma-glutamylcysteine ligase [GCLC (rate-limiting enzyme)], were quantified. The hepatic GCLC transcript and protein declined in Allo compared to Syn mice [GCLC fold change: day +4 = 0.4 vs 1.03±0.15; day +10 = 0.3± 0.05 vs 1.0± 0.05 (p = 0.0004), respectively]. Target organs like lungs and gut also revealed depletion of intracellular thiol in Allo mice that directly correlated with the degree of T cell infiltration. Together, these data show that the GSH synthetic pathway is specifically affected in GVHD. The tight transcriptional deregulation of GCLC with depletion of GSH reserve is an early event and precedes histopathological changes of GVHD. Identification of modifiers of GCLC/GSH/GSSG system may provide a novel target for treatment of GVHD.

Oxidative stress in Cyprinus carpio to analyze microcystin impact in eutrophic shallow lakes: a preliminary study

Microcystins (MCYSTs) are toxins produced by cyanobacteria in aquatic environment and are of high potential risk to aquatic organisms. The physiological responses and pathobiological developments that they elicit in fish have been extensively studied, mainly through acute toxicity experiments. This study was designed to examine the seasonal fluctuation of biochemical markers of oxidative stress in different tissues of a natural population of Cyprinus carpio inhabiting a shallow Mediterranean lake, along with the respective MCYSTs concentrations in blood and tissues at environmentally relevant MCYSTs values. MCYSTs content was assessed in liver, kidney, intestine, brain and muscle along with the MCYSTs in lake water and scum applying ELISA technique. Catalase activity, GSH/GSSH relative concentrations and lipid peroxidation were used as biochemical markers. Our results suggest that common carp of Lake Pamvotis exposed to naturally fluctuating concentrations of MCYST in water and scum contained stably high MCYST concentrations in all tissues that might pose a threat to public health. Liver and kidney were the primary target organs. Tissue concentrations did not correlate with the response of any of the elements of the antioxidant defence system. Hepatic catalase, GSH content and TBARS in all tissues tested followed the fluctuations of major limnological parameters, i.e. water temperature and oxygen concentration, chlorophyll-a, MCYST in water and scum, suggesting that they should be cautiously used to monitor exposure to MCYSTS in natural freshwater ecosystems.

THE PROTECTIVE EFFECT OF COFFEE AGAINST PARACETAMOL-INDUCED HEPATIC INJURY IN RATS

ABSTRACT The effect of daily coffee ingestion on the hepatic detoxifying enzymes of rats challenged with high doses of paracetamol (PCM) was investigated. Two groups of male Wistar rats fed, respectively, chow diets supplemented or not with a coffee extract, were sacrificed at 75 days of age, 48 h after injection of saline or PCM (700 mg/kg body wt). The hepatocytes were isolated. Coffee decreased the lipid peroxidation (29%), increased glutathione (GSH) content (46%) and abolished the increased peroxidation caused by PCM (25%). PCM reduced the hepatic GSH content (62%), but in the animals fed the coffee supplemented diet, the reduction was less pronounced (32%). Coffee increased liver UDP-glucuronosyl-transferases UGT1 (75%) and UGT2 (38%), glutathione transferase (46%) and glutathione reductase (72%) activities and decreased glutathione peroxidase (31%). Therefore, it can be concluded that coffee ingestion modulates phase II of the hepatic biotransformation system improving the detoxification ability of the organism against toxic xenobiotics. PRACTICAL APPLICATIONS Coffee is a very popular beverage, which is consumed worldwide and consists of many substances that are suggested to be potential chemoprotectors in biological systems. Among the constituents, caffeine, phenolic polymers, chlorogenic acids, lipids, kahweol and cafestol, all of which have pharmacological properties that lead to adaptation of the organism to its environment, can be cited. Coffee, therefore, is able to protect the human being against diseases and xenobiotic challenges. The results presented in this study demonstrate the effect of coffee as a modulator of the hepatic detoxifying enzyme system. Therefore, this beverage can be classified as a functional food. In addition to the reported application of coffee for the protection against degenerative diseases, free radical damage, cancer and other metabolic alterations, it also acts by modulating the hepatic enzyme system, thereby improving the detoxification capacity of the liver.

Effectiveness of Melatonin, as a Radiation Damage-Mitigating Drug in Modulating Liver Biochemical disorders in γ-Irradiated Rats

MELATONIN has an antiperoxidative effect on several tissues as well as a scavenger effect on reactive oxygen species (ROS). Whilst radiation-hazards due to free radical generation, present enormous challenges for biological and medical safety. Therefore, rats were classified into four groups; control (n= 8), (received 0.5 ml of alchoholic saline as a vehicle for 5 days). Melatonin-treated rats received 10 mg/ kg body wt, for 5 days (given to the animals in the morning via stomach tube). γ-irradiated rats received 0.5 ml of the melatonin vehicle followed by one shot dose of 3 Gy γ-rays. Each of these groups was compared with a further group, which-received melatonin for 5 days after 3 Gy γ-irradiation exposure. The results revealed that all considered biochemical parameters were not changed significantly in melatonin-treated group as compared with control one. In the liver tissue of the γ-irradiated animals (3 Gy), the oxidative stress markers malondialdehyde (MDA) and protein carbonyl (PC) were significantly increased, while a marked decrease occurred in the contents of deoxy- & ribo-nucleic acids (DNA & RNA) and glutathione (GSH) as well as activity of glutathione-S-transferase (GST). In addition, catalase (CAT) and myeloperoxidase (MPO) activities were increased. Activities of aspartate transaminase (AST), alkaline phosphatase (ALP) and gamma-glutamyltransferase (GGT) were significantly increased in sera of the irradiated rats. Treatment with melatonin for 5 days after γ-rays exposure significantly modulated the radiation-induced elevations in MDA and PC levels in the liver tissue and significantly restored hepatic GSH content, GST, CAT and MPO activities. Post-irradiation treatment with melatonin showed significant higher hepatic DNA and RNA contents than irradiated rats. The activities of AST, ALP, and GGT in serum were significantly ameliorated when melatonin was administrated after irradiation.

Protective Mechanism of Andrographolide against Carbon Tetrachloride-Induced Acute Liver Injury in Mice

The aim of this study was to investigate the protective effects of andrographolide (AP), a bioactive component isolated from Andrographis paniculata, on carbon tetrachloride (CCl(4))-induced liver injury as well as the possible mechanisms involved in this protection in mice. Acute liver injury was induced by CCl(4) intoxication in mice. Serum biological analysis, lipid peroxides and antioxidant estimation, histopathological studies, reverse transcription polymerase chain reaction (RT-PCR) and Western blot assay were carried out. CCl(4) treatment resulted in severe hepatic injury, as evidenced by significant elevation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels and typical histopathological changes, such as hepatocyte necrosis. Additionally, CCl(4) administration led to oxidative stress in mice, as indicated by a remarkable increase in the hepatic malondialdehyde (MDA) level, together with a significant decrease in liver reduced glutathione (GSH) content. However, CCl(4)-induced hepatotoxicity was significantly attenuated by pretreatment with AP, as demonstrated by significant reduction of serum ALT, AST levels and hepatic MDA activity, along with a remarkable increase in hepatic GSH content. Histopathological changes induced by CCl(4) were also ameliorated by AP pretreatment. The marked increase of tumor necrosis factor-α (TNF-α) induced by CCl(4) was attenuated by AP, and the dramatic elevation of heme oxygenase-1 (HO-1) at transcriptional and protein levels was augmented following AP pretreatment. AP can effectively prevent liver injury induced by CCl(4), which may be due to inhibition of oxidative stress and inflammatory responses.