Glutathione S-transferase Enzyme Activity Research Articles

The colonization of pyrethroid resistant strain from wild Anopheles sinensis, the major Asian malaria vector.

BackgroundAnopheles sinensis is one of the most important malaria vectors in Asian countries. The rapid spread of insecticide resistance has become a major obstacle for insecticide-based strategies for vector control. Therefore, it is necessary to prepare an insecticide-resistant strain of An. sinensis to further understand the insecticide resistance mechanisms in this species to facilitate genetic approaches to targeting the insecticide-resistant population of this important malaria vector.MethodsAn. sinensis mosquitoes were collected from regions where pyrethroid resistance had been reported. The mosquitoes were subjected to continuous pyrethroid selection after species confirmation, and the forced copulation method was used to increase the mating rate. In addition, the knockdown-resistance (kdr) mutation frequencies of each generation of An. sinensis were measured; and the metabolic enzyme activities of cytochrome P450 monoxygenases (P450s) and glutathione S-transferases (GSTs) were detected.ResultsThe identification of field-captured An. sinensis was confirmed by both morphological and molecular methods. The population of An. sinensis exhibited stable resistance to pyrethroid after continuous generations of pyrethroid selection in the laboratory with high kdr mutation frequencies; and elevated levels of both P450s and GSTs were significantly found in field selected populations comparing with the laboratory susceptible strain. So far, the colonised strain has reached its eleventh generation and culturing well in the laboratory.ConclusionsWe colonised a pyrethroid-resistant population of An. sinensis in the laboratory, which provides a fundamental model for genetic studies of this important malaria vector.

The potential medicinal value of plants from Asteraceae family with antioxidant defense enzymes as biological targets

Context: Plants and most of the plant-derived compounds have long been known for their potential pharmaceutical effects. They are well known to play an important role in the treatment of several diseases from diabetes to various types of cancers. Today most of the clinically effective pharmaceuticals are developed from plant-derived ancestors in the history of medicine.Objective: The aim of this study was to evaluate the free radical scavenging activity and total phenolic and flavonoid contents of methanol, ethanol, and acetone extracts from flowers and leaves of Onopordum acanthium L., Carduus acanthoides L., Cirsium arvense (L.) Scop., and Centaurea solstitialis L., all from the Asteraceae family, for investigating their potential medicinal values of biological targets that are participating in the antioxidant defense system such as catalase (CAT), glutathione S-transferase (GST), and glutathione peroxidase (GPx).Materials and methods: In this study, free radical scavenging activity and total phenolic and flavonoid contents of the plant samples were assayed by DPPH, Folin–Ciocalteu, and aluminum chloride colorimetric methods. Also, the effects of extracts on CAT, GST, and GPx enzyme activities were investigated.Results and discussion: The highest phenolic and flavonoid contents were detected in the acetone extract of C. acanthoides flowers, with 90.305 mg GAE/L and 185.43 mg Q/L values, respectively. The highest DPPH radical scavenging was observed with the methanol leaf extracts of C. arvense with an IC50 value of 366 ng/mL. The maximum GPx and GST enzyme inhibition activities were observed with acetone extracts from the flower of C. solstitialis with IC50 values of 79 and 232 ng/mL, respectively.

Evaluation of zinc oxide nanoparticle toxicity in sludge products applied to agricultural soil using multispecies soil systems

To study the environmental impact of nanoparticles, the sludges of wastewater (WWTS) and water treatment (WTS) plants enriched with ZnO nanoparticles were added to agricultural soil, and the toxic effects of the nanoparticles were studied using a microcosm system based on the soil. The WWTS treated soils were characterised by statistically significant decreases (p<0.05) in Vicia sativa germination at the lowest (76.2%) and medium (95.2%) application rates, decreases in the fresh biomass for Triticum aestivum (19.5%), Raphanus sativus (64.1%), V. sativa (37.4%) and Eisenia fetida (33.6%) at the highest application rate and a dose-related significant increase (p<0.05) in earthworm mortality. In WTS amended soils, significant reductions (p<0.05) of the fresh biomass (17.2%) and the chlorophyll index (24.4%) for T. aestivum and the fresh biomass for R. sativus (31.4%) were only recorded at the highest application doses. In addition, the soil phosphatase enzymatic activity decreased significantly (p<0.05) in both WWTS (dose related) and WTS treatments. For water organisms, a slight inhibition of the growth of Chlorella vulgaris was observed (WWTS treated soils), along with statistically significant dose-related inhibition responses on total glutathione cell content, and statistically significant dose-related induction responses on the glutathione S-transferase enzyme activity and the reactive oxygen species generation on the RTG-2 fish cell line.

Evaluation of gatifloxacin for its potential to induce antioxidant imbalance and retinopathy in rabbits

Gatifloxacin, a fluoroquinolone antibiotic, has been reported to produce several adverse reactions. In the present investigation, gatifloxacin administered at the dose rate of 10 and 20 mg kg(-1) body weight per day, respectively, for 21 consecutive days, was evaluated for its potential to induce antioxidant status alterations and retinal damage in rabbits. A significant alteration in the antioxidant status of rabbits particularly in the high-dose group was observed which is indicated by decreased activity of superoxide dismutase and levels of blood glutathione with a concomitant increase in the activity of catalase, glutathione peroxidase, and glutathione S-transferase enzymes. The activity of glutathione reductase differed nonsignificantly between groups throughout the study period. The levels of malondialdehyde were elevated in the high-dose group. The histopathological examination of eyeball tunics revealed clumping of nuclei of the retinal outer nuclear layer in the gatifloxacin-treated groups. The results from this study indicate that repeated gatifloxacin administration produces a dose-dependent oxidative stress and retinopathy.

Applicative implications of Carcinus maenas and Ruditapes philippinarum in biomonitoring studies after oil spills

Biomarkers have been tested in order to address the most suitable battery for determining adverse effects of crude oil spills on marine invertebrates. An oil spill with increasing degrees of severity was simulated by mixing crude oil (0%, 0.5%, 2%, 8%, 16%, 32%) with sediment. Carcinus maenas and Ruditapes philippinarum were exposed to this sediment for seven days with the aim of comparing their applicability in biomonitoring studies. Four biomarkers including ethoxyresorufin O-deethylase (EROD), glutathione S-transferase (GST), glutathione peroxidase (GPx) and lipid peroxidation (LPO) were analysed in gill and digestive gland tissues of clams; and in gill and hepato-pancreas tissues of crabs. EROD, GST and GPx enzymatic activities were significantly induced in gill and digestive gland tissues of clams when increasing oil concentrations (p<.01). In crabs all the biomarkers were significantly activated in gill tissues, whereas EROD and LPO activities were induced only in hepato-pancreas tissues (p<.01). Gill and digestive gland in clams and gill in crabs were found to be the most reliable tissues for analysis of biomarkers. The biomarkers selected are thus considered suitable for assessing toxicity of sediments after a marine crude oil spill accident. Both species were found to be sensitive and suitable for biomonitoring purposes.

Inhibition of the recombinant cattle tick Rhipicephalus (Boophilus) annulatus glutathione S-transferase

Rhipicephalus (Boophilus) annulatus is a bloodsucking ectoparasite that causes severe production losses in the cattle industry. This study aims to evaluate the in vitro effects of tannic acid, hematin (GST inhibitors) and different plant extracts (rich in tannic acid) on the activity of the recombinant glutathione S-transferase enzyme of the Egyptian cattle tick R. annulatus (rRaGST), in order to confirm their ability to inhibit the parasitic essential detoxification enzyme glutathione S-transferase. Extraction with 70% ethanol of Hibiscus cannabinus (kenaf flowers), Punica granatum (red and white pomegranate peel), Musa acuminata (banana peel) (Musaceae), Medicago sativa (alfalfa seeds), Tamarindus indicus (seed) and Cuminum cyminum (cumin seed) were used to assess: (i) inhibitory capacities of rRaGST and (ii) their phenolic and flavonoid contents. Ethanol extraction of red pomegranate peel contained the highest content of phenolic compounds (29.95mg gallic acid/g dry tissue) compared to the other studied plant extracts. The highest inhibition activities of rRaGST were obtained with kenaf and red pomegranate peel (P. granatum) extracts with IC50 values of 0.123 and 0.136mg dry tissue/ml, respectively. Tannic acid was the more effective inhibitor of rRaGST with an IC50 value equal to 4.57μM compared to delphinidine-HCl (IC50=14.9±3.1μM). Gossypol had a weak inhibitory effect (IC50=43.7μM), and caffeic acid had almost no effect on tick GST activity. The IC50 values qualify ethacrynic acid as a potent inhibitor of rRaGST activity (IC50=0.034μM). Cibacron blue and hematin showed a considerable inhibition effect on rRaGST activity, and their IC50 values were 0.13μM and 7.5μM, respectively. The activity of rRaGST was highest for CDNB (30.2μmol/min/mg protein). The enzyme had also a peroxidatic activity (the specific activity equals 26.5μmol/min/mg protein). Both tannic acid and hematin inhibited rRaGST activity non-competitively with respect to GSH and competitively with respect to CDNB. While red pomegranate extracts inhibited rRaGST activity competitively with respect to GSH, uncompetitive inhibition was observed with respect to CDNB.

Effects of UV radiation on hatching, lipid peroxidation, and fatty acid composition in the copepod Paracyclopina nana

To evaluate the effects of UV radiation on the reproductive physiology and macromolecules in marine zooplankton, several doses of UV radiation were used to treat the copepod Paracyclopina nana, and we analyzed in vivo endpoints of their life cycle such as mortality and reproductive parameters with in vitro biochemical biomarkers such as reactive oxygen species (ROS), the modulated enzyme activity of glutathione S-transferase (GST) and superoxide dismutase (SOD), and the production of a byproduct of peroxidation (e.g. malonedialdehyde, MDA). After UV radiation, the survival rate of P. nana was significantly reduced. Also, egg sac damage and a reduction in the hatching rate of offspring were observed in UV-irradiated ovigerous females. According to the assessed biochemical parameters, we found dose-dependent increases in ROS levels and high levels of the lipid peroxidation decomposition product by 2kJm−2, implying that P. nana was under off-balanced status by oxidative stress-mediated cellular damage. Antioxidant enzyme activities of GST and SOD increased over different doses of UV radiation. To measure UV-induced lipid peroxidation, we found a slight reduction in the composition of essential fatty acids such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). These findings indicate that UV radiation can induce oxidative stress-triggered lipid peroxidation with modulation of antioxidant enzyme activity, leading to a significant effect on mortality and reproductive physiology (e.g. fecundity). These results demonstrate the involvement of UV radiation on essential fatty acids and its susceptibility to UV radiation in the copepod P. nana compared to other species.

Antioxidative activity of different parts of the plant Lepidium sativum Linn

Lepidium sativum Linn. has been used in traditional and folklore medicine for the treatment of bronchial asthma, diabetes, local and rheumatic pain. An ethanolic extract of cress (L. sativum L.) shoot, leaf, stem and seed has been studied for antioxidative active against 1,1-diphenyl-2-picrylhydrazyl (DPPH), total glutathione S-transferase assay, reduced glutathione activity, reducing power (Fe3+–Fe2+ Transformation Ability), and ascorbic acid is also estimated. The percentage yields of free radical scavenging activity (DPPH) obtained for different ethanolic extracts of L. sativum. Supreme scavenging activity was detected in shoot (12.19±02%) and least in stem (2.69±05%). The activity of total glutathione S-transferase enzyme was found to be more in seed (9600±56.3μg/ml) than other plant parts. The reduced glutathione content of the ethanolic extracts of L. sativum was found to be more in leaf (9±0.2μg/ml). In the reducing power assay, ethanolic extracts gives the optical density in increasing concentration in all plant parts it shows that it has the reducing ability of Fe3+–Fe2+. Presence of vitamin C was tested. It was found that the shoot extract has highest amount of vitamin C. The results of present data were shown that the ethanolic extract of L. sativum L. plant parts have contributed high potential in vitro antioxidant activity.

Perinatal manganese exposure and hydroxyl radical formation in rat brain.

The present study was designed to investigate the role of pre- and postnatal manganese (Mn) exposure on hydroxyl radical (HO•) formation in the brains of dopamine (DA) partially denervated rats (Parkinsonian rats). Wistar rats were given tap water containing 10,000 ppm manganese chloride during the duration of pregnancy and until the time of weaning. Control rat dams consumed tap water without added Mn. Three days after birth, rats of both groups were treated with 6-hydroxydopamine at one of three doses (15, 30, or 67 µg, intraventricular on each side), or saline vehicle. We found that Mn content in the brain, kidney, liver, and bone was significantly elevated in dams exposed to Mn during pregnancy. In neonates, the major organs that accumulated Mn were the femoral bone and liver. However, Mn was not elevated in tissues in adulthood. To determine the possible effect on generation of the reactive species, HO• in Mn-induced neurotoxicity, we analyzed the contents of 2.3- and 2.5-dihydroxybenzoic acid (spin trap products of salicylate; HO• being an index of in vivo HO• generation), as well as antioxidant enzyme activities of superoxide dismutase (SOD) isoenzymes and glutathione S-transferase (GST). 6-OHDA-depletion of DA produced enhanced HO• formation in the brain tissue of newborn and adulthood rats that had been exposed to Mn, and the latter effect did not depend on the extent of DA denervation. Additionally, the extraneuronal, microdialysate, content of HO• in neostriatum was likewise elevated in 6-OHDA-lesioned rats. Interestingly, there was no difference in extraneuronal HO• formation in the neostriatum of Mn-exposed versus control rats. In summary, findings in this study indicate that Mn crosses the placenta but in contrast to other heavy metals, Mn is not deposited long term in tissues. Also, damage to the dopaminergic system acts as a “trigger mechanism,” initiating a cascade of adverse events leading to a protracted increase in HO• generation, and the effects of Mn and 6-OHDA are compounded. Moreover, HO• generation parallels the suppression of SOD isoenzymes and GST in the brains of rats lesioned with 6-OHDA and/or intoxicated with Mn—the most prominent impairments being in frontal cortex, striatum, and brain stem. In conclusion, ontogenetic Mn exposure, resulting in reactive oxygen species, HO• formation, represents a risk factor for dopaminergic neurotoxicity and development of neurodegenerative disorders.

C9,t11-Conjugated linoleic acid ameliorates steatosis by modulating mitochondrial uncoupling and Nrf2 pathway

Oxidative stress, hepatic steatosis, and mitochondrial dysfunction are key pathophysiological features of nonalcoholic fatty liver disease. A conjugated linoleic acid (CLA) mixture of cis9,trans11 (9,11-CLA) and trans10,cis12 (10,12-CLA) isomers enhanced the antioxidant/detoxifying mechanism via the activation of nuclear factor E2-related factor-2 (Nrf2) and improved mitochondrial function, but less is known about the actions of specific isomers. The differential ability of individual CLA isomers to modulate these pathways was explored in Wistar rats fed for 4 weeks with a lard-based high-fat diet (L) or with control diet (CD), and, within each dietary treatment, two subgroups were daily administered with 9,11-CLA or 10,12-CLA (30 mg/day). The 9,11-CLA, but not 10,12-CLA, supplementation to CD rats improves the GSH/GSSG ratio in the liver, mitochondrial functions, and Nrf2 activity. Histological examination reveals a reduction of steatosis in L-fed rats supplemented with both CLA isomers, but 9,11-CLA downregulated plasma concentrations of proinflammatory markers, mitochondrial dysfunction, and oxidative stress markers in liver more efficiently than in 10,12-CLA treatment. The present study demonstrates the higher protective effect of 9,11-CLA against diet-induced pro-oxidant and proinflammatory signs and suggests that these effects are determined, at least in part, by its ability to activate the Nrf2 pathway and to improve the mitochondrial functioning and biogenesis.

Effect of mixture of diazinon and benzo[a]pyrene in Glutathione S-transferase of Nile tilapia

Currently, with the growing contamination of aquatic ecosystems, many compounds, such as pesticides and hydrocarbons, are improperly released into rivers and lakes. Studies about the exposure of single pollutants causing biochemical variations are abundant in the literature. However, there are few studies focused on the biological effects of complex mixtures. The aim of this investigation was to evaluate whether mixture diazinon and benzo[a]pyrene can affect the biochemical activities of classic biomarkers such as acetylcholinesterase (AChE), carboxylesterase (CbE), catalase (CAT), glutathione peroxidase (GPx) and glutathione S-transferase (GST) in Nile tilapia and compare the effects on enzymatic systems of the exposure to a mixture of compounds and the effects observed when they are exposed separately. We measured the activity of enzymes in gills and liver of Nile Tilapia (Oreochromis niloticus) after 2 and 7 days of exposure to Diazinon (0.5 mg/L) and benzo[a]pyrene (1.0 mg/L) individually and in mixture. The results showed that in the mixture group after 7 days of exposure, the benzo[a]pyrene increased the inhibitory action of Diazinon in GST enzyme activity, in liver tissue. This indicates that the toxicity of the interactions between pesticide and polycyclic aromatic hydrocarbon may show synergistic effect. These results suggest the importance of studies with mixture of compounds, because these data will help to understand the results obtained in field studies, such as those from environmental monitoring.

The colonization of pyrethroid resistant strain from wild Anopheles sinensis , the major Asian malaria vector

BackgroundAnopheles sinensis is one of the most important malaria vectors in Asian countries. The rapid spread of insecticide resistance has become a major obstacle for insecticide-based strategies for vector control. Therefore, it is necessary to prepare an insecticide-resistant strain of An. sinensis to further understand the insecticide resistance mechanisms in this species to facilitate genetic approaches to targeting the insecticide-resistant population of this important malaria vector.MethodsAn. sinensis mosquitoes were collected from regions where pyrethroid resistance had been reported. The mosquitoes were subjected to continuous pyrethroid selection after species confirmation, and the forced copulation method was used to increase the mating rate. In addition, the knockdown-resistance (kdr) mutation frequencies of each generation of An. sinensis were measured; and the metabolic enzyme activities of cytochrome P450 monoxygenases (P450s) and glutathione S-transferases (GSTs) were detected.ResultsThe identification of field-captured An. sinensis was confirmed by both morphological and molecular methods. The population of An. sinensis exhibited stable resistance to pyrethroid after continuous generations of pyrethroid selection in the laboratory with high kdr mutation frequencies; and elevated levels of both P450s and GSTs were significantly found in field selected populations comparing with the laboratory susceptible strain. So far, the colonised strain has reached its eleventh generation and culturing well in the laboratory.ConclusionsWe colonised a pyrethroid-resistant population of An. sinensis in the laboratory, which provides a fundamental model for genetic studies of this important malaria vector.

Resistance selection with cadmium and changes in the activities of antioxidases in Boettcherisca peregrina (Diptera: Sarcophagidae)

In order to establish a physiological link between antioxidases and the resistance level of insects to cadmium (Cd), natural populations of Boettcherisca peregrina (Diptera: Sarcophagidae) were maintained for 20 generations and reared either on an uncontaminated diet or on a diet contaminated with cadmium (Cd) at a concentration equivalent to the median lethal concentration (LC50) as determined every five generations. A relatively susceptible strain (S) and a Cd-resistant strain (R) were selected. The metal accumulation, growth and development, reproduction, and antioxidant enzyme activities in these strains were analyzed. The results showed that R-strain organisms had enhanced juvenile survivorship, increased Cd accumulation, and increased adult female fecundity when compared with S-strain. The larval enzyme activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and glutathione S-transferase (GST) in R-strain larvae were higher than those in S-strain larvae when fed diets with or without Cd. This indicates that Cd resistance in B. peregrina larvae is mediated by SOD, CAT, GR, and GST.

Effect of pectolinaringenin, a flavonoid from Clerodendrum phlomidis, on total protein, glutathione S-transferase and esterase activities of Earias vittella and Helicoverpa armigera

Pectolinaringenin was isolated from chloroform extract of Clerodendrum phlomidis L. and was evaluated at 12.5 to 100 ppm concentrations for its effects on total protein, esterase and glutathione S-transferase (GST) activities of Earias vittella and Helicoverpa armigera. At 100 ppm, the compound reduced total protein content by 55.75% and 53.01% over control with IC50 values of 74.37 and 212.31 ppm in E. vittella and H. armigera, respectively. At 100 ppm it also reduced GST and esterase enzyme activities in E. vittella by 37.53% and 43.09% over control, with IC50 values of 133.00 and 111.76 ppm, respectively. It also reduced GST and esterase activities in H. armigera by 43.14% and 47.421% over control with IC50 values of 114.38 and 98.78 ppm, respectively. Data were analyzed for their normality using the Shapiro-Wilk test and Levene’s statistics to determine the significant deviation. Pectolinaringenin could be used for the management of agricultural pests.

Functional compensation of glutathione S-transferase M1 (GSTM1) null by another GST superfamily member, GSTM2

The gene for glutathione-S-transferase (GST) M1 (GSTM1), a member of the GST-superfamily, is widely studied in cancer risk with regard to the homozygous deletion of the gene (GSTM1 null), leading to a lack of corresponding enzymatic activity. Many of these studies have reported inconsistent findings regarding its association with cancer risk. Therefore, we employed in silico, in vitro, and in vivo approaches to investigate whether the absence of a functional GSTM1 enzyme in a null variant can be compensated for by other family members. Through the in silico approach, we identified maximum structural homology between GSTM1 and GSTM2. Total plasma GST enzymatic activity was similar in recruited individuals, irrespective of their GSTM1 genotype (positive/null). Furthermore, expression profiling using real-time PCR, western blotting, and GSTM2 overexpression following transient knockdown of GSTM1 in HeLa cells confirmed that the absence of GSTM1 activity can be compensated for by the overexpression of GSTM2.

Effects of norfloxacin on the drug metabolism enzymes of two sturgeon species (Acipenser schrenckiandAcipenser ruthenus)

Summary The enzyme activities of 7-ethoxycoumarin O-deethylase (ECOD) and glutathione S-transferase (GST), and on glutathione (GSH) content, involved in metabolism of the antibiotic Norfloxacin (NFLX), were investigated in Acipenser schrencki and Acipenser ruthenus. Sturgeons weighing 45–55 g were kept in an aquarium (0.5 m × 0.5 m × 0.9 m) for two weeks under controlled conditions (fish density 88 individuals per m3, 18°C) before the experiment. The two species of sturgeon were divided into five groups each (n = 15 in each group), with each group subdivided into three replicates of five fish per tank. A control group in which distilled water was administered orally was also tested. NFLX was forced into the stomachs of the fish at a concentration of 20, 40, 60, 80 and 100 mg kg−1 body weight, respectively. ECOD activities in liver microsomes, and GST activity and GSH content in liver microsomes and blood plasma, were measured and compared. Results indicate that ECOD activity is progressively inhibited with increasing NFLX concentrations. ECOD activity varied from 0.12 nmol mg−1 min−1 to 0.07 nmol mg−1 min−1, demonstrating an inhibition rate of 60.83% in A. schrencki and 65.14% in A. ruthenus. In both species tested, GST and GSH levels exhibited a trend of first increasing, and then decreasing with increasing NFLX levels, reaching a peak value at 40 mg per kg−1 body weight. Thus, the presented results indicate that NFLX can induce a change in the activity of some drug metabolism related enzymes such as ECOD and GST in vivo.

Trabectedin to induce mitochondrial membrane potential dissipation and reactive oxygen species generation in breast cancer cells.

e13580 Background: Trabectedin (Yondelis; ET-743) is a tetrahydroisoquinoline alkaloid that was originally derived from the marine tunicate Ecteinascidia turbinata and is currently prepared synthetically. It ıs currently used for the treatment of soft tissue sarcomas. It has been shown that Trabectedin provides the production of superoxides near the DNA strand, resulting in DNA backbone cleavage and cell apoptosis, however the apoptotic mechanisms of Trabectedin is still very limited. The objective of this study is to investigate the underlying mechanisms of apoptosis Trabectedin in two human breast cancer cell lines (MDA-MB-231 and MDA-MB-453) and one human immortalized non-transformed breast epithelial cell line (MCF-10A), to see if similar oxidation processes take place in breast cancer. Methods: Cytotoxicity was assessed by XTT cell viability assay. Apoptosis was shown by measuring DNA fragmentation, caspase 3/7 activation. Mitochondrial membrane potential (MMP) was evaluated by TMRE dye. Measurement of reactive oxygen species (ROS) was done by using Glutathione S-Transferase (GST) Assay Kit and CM-H2DCFDA dye. Results: Trabectedin induced the cytotoxicity in breast cancer cells in a time and dose dependent manner. Moreover, it increased DNA fragmentation and the MMP dissipation in tested breast cancer cells. The levels of ROS production in parallel with GST enzyme activity were sharply increased by Trabectedin treatment. Conclusions: This is the first study to investigate the role of Trabectedin activity and mechanisms of apoptosis in human breast cancer cells. These preliminary results might show us a way to use Trabectedin alone, or in combination for breast cancer treatment in near future.

Glutathione S-transferase (GST) family in barley: Identification of members, enzyme activity, and gene expression pattern

Barley (Hordeum vulgare) is one of the most important cereals in many developing countries where drought stress considerably diminishes agricultural production. Glutathione S-transferases (GSTs EC 2.5.1.18) are multifunctional enzymes which play a crucial role in cellular detoxification and oxidative stress tolerance. In this study, 84 GST genes were identified in barley by a comprehensive in silico approach. Sequence alignment and phylogenetic analysis grouped these HvGST proteins in eight classes. The largest numbers of the HvGST genes (50) were included in the Tau class followed by 21 genes in Phi, five in Zeta, two in DHAR, two in EF1G, two in Lambda, and one each in TCHQD and Theta classes. Phylogenetic analysis of the putative GSTs from Arabidopsis, rice, and barley indicated that major functional diversification within the GST family predated the monocot/dicot divergence. However, intra-specious duplication seems to be common. Expression patterns of five GST genes from Phi and Tau classes were investigated in three barley genotypes (Yusof [drought-tolerant], Moroc9-75 [drought-sensitive], and HS1 [wild ecotype]) under control and drought-stressed conditions, during the vegetative stage. All investigated genes were up-regulated significantly under drought stress and/or showed a higher level of transcripts in the tolerant cultivar. Additionally, GST enzyme activity was superior in Yusof and induced in the extreme-drought-treated leaves, while it was not changed in Moroc9-75 under drought conditions. Moreover, the lowest and highest levels of lipid peroxidation were observed in the Yusof and Moroc9-75 cultivars, respectively. Based on the achieved results, detoxification and antioxidant activity of GSTs might be considered an important factor in the drought tolerance of barley genotypes for further investigations.

The Effect of Date Palm Fruit (Phoenix dactylifera L.) on the Enzyme Glutathione-S-Transferase Activity in Sprague-Dawley Rats

This research was performed to study the effect of date palm fruit (Phoenix dactylifera L.) on the hepatic enzyme system glutathione-S-transferase (GST) in rats with 7, 12-dimethylbenz (alpha) anthracene (DMBA)-induced mammary cancer. The effect of feeding the fruit was compared to that of feeding raw soybean seeds and injection with 2 drugs (preventive and curative against mammary cancer) i.e., tamoxifien and 17-beta-estradiol. Eighty three female weanling Sprague-Dawley rats were randomly distributed into 8 groups. Each group received one of the dietary or drug treatments. One of the groups was injected with sesame oil which is the carcinogen vehicle, to serve as a negative/DMBA control group. Another group of rats that received neither drug nor special dietary treatment served as a positive/DMBA control group. The experiment lasted for 26 weeks. At the end of the feeding period, all animals were killed, livers were removed, prepared and analyzed for their protein content and GST enzyme activity. Livers of the rats that were injected with sesame oil vehicle showed the highest enzyme activity. The rats which were fed the date palm fruit tended to (p = 0.079) reduce the enzyme activity. It is concluded that the date palm fruit at the 2 maturity stages may possess antioxidant activity that is reflected positively in the prevention of DMBA-induced mammary cancer.

Identification of Allergenic Component Tyr p 8 from Tyrophagus putrescentiae and Cross-Reactivity with Der p 8

Group 8 mite allergens exhibit sequence homology to glutathione S-transferases (GSTs), such as that from Dermatophagoides pteronyssinus (Der p 8). GSTs have been identified as important allergens in studies of allergens from house dust mites, cockroaches, and fungi. Our objective was to purify the native group 8 allergen from Tyrophagus putrescentiae (nTyr p 8) and generate recombinant Tyr p 8 (rTyr p 8) for immunological characterization. The allergenicity was determined by antibody recognition, IgE inhibition, and triggering of the basophil-sensitized release of histamine, using T. putrescentiae hypersensitivity sera. The results showed that the mRNA transcript of nTyr p 8 is 657 bp long, contains 218 amino acids with a molecular mass of 26 kDa, and exhibits 83% sequence homology to Der p 8. Serum samples from the allergic patients with an IgE-positive response to T. putrescentiae were analyzed to determine their IgE response to rTyr p 8. The results showed that the sera of 48 subjects (45.3%) had specific IgE against rTyr p 8. However, sera of only 19 subjects (17.9%) had specific IgE against rTyr p 8 after D. pteronyssinus absorption. Histamine release was observed from T. putrescentiae-allergic subjects in the presence of rTyr p 8. Both the nTyr p 8 and T. putrescentiae crude extract had been demonstrated to possess GST enzymatic activity. Although the specific binding of serum IgE to rTyr p 8 was only 17.9%, which indicates that rTyr p 8 was not a major allergen, the positive response to rTyr p 8 was due to the cross-reactivity with Der p 8. The group 8 mite allergen might be of use in the design of a suitable allergen for diagnosis and for the development of novel immunotherapies.