Novel aryl beta-aminocarbonyl derivatives as inhibitors of Trypanosoma cruzi trypanothione reductase: binding mode revised by docking and GRIND2-based 3D-QSAR procedures

Trypanothione reductase has long been investigated as a promising target for chemotherapeutic intervention in Chagas disease, since it is an enzyme of a unique metabolic pathway that is exclusively present in the pathogen but not in the human host, which has the analog Glutathione reductase. In spite of the present data-set includes a small number of compounds, a combined use of flexible docking, pharmacophore perception, ligand binding site prediction, and Grid-Independent Descriptors GRIND2-based 3D-Quantitative Structure-Activity Relationships (QSAR) procedures allowed us to rationalize the different biological activities of a series of 11 aryl beta-aminocarbonyl derivatives, which are inhibitors of Trypanosoma cruzi trypanothione reductase (TcTR). Three QSAR models were built and validated using different alignments, which are based on docking with the TcTR crystal structure, pharmacophore, and molecular interaction fields. The high statistical significance of the models thus obtained assures the robustness of this second generation of GRIND descriptors here used, which were able to detect the most important residues of such enzyme for binding the aryl beta-aminocarbonyl derivatives, besides to rationalize distances among them. Finally, a revised binding mode has been proposed for our inhibitors and independently supported by the different methodologies here used, allowing further optimization of the lead compounds with such combined structure- and ligand-based approaches in the fight against the Chagas disease.

Inhibitors of Trypanosoma brucei trypanothione reductase: comparative molecular field analysis modeling and structural basis for selective inhibition

Background: Sleeping sickness is a major cause of death in Africa. Since no secure treatment is available, the development of novel therapeutic agents is urgent. In this context, the enzyme trypanothione reductase (TR) is a prominent molecular target that has been investigated in drug design for sleeping sickness. Results: In this study, comparative molecular field analysis models were generated for a series of Trypanosoma brucei TR inhibitors. Statistically significant results were obtained and the models were applied to predict the activity of external test sets, with good correlation between predicted and experimental results. We have also investigated the structural requirements for the selective inhibition of the parasite's enzyme over the human glutathione reductase. Conclusion: The quantitative structure-activity relationship models provided valuable information regarding the essential molecular requirements for the inhibitory activity upon the target protein, providing important insights into the design of more potent and selective TR inhibitors.

Fructose-1,6-bisphosphate preserves intracellular glutathione and protects cortical neurons against oxidative stress

Fructose-1,6-bisphosphate (FBP), an endogenous intermediate of glycolysis, protects the brain against ischemia-reperfusion injury. The mechanisms of FBP protection after cerebral ischemia are not well understood. The current study was undertaken to determine whether FBP protects primary neurons against hypoxia and oxidative stress by preserving reduced glutathione (GSH). Cultures of pure cortical neurons were subjected to oxygen deprivation, a donor of nitric oxide and superoxide radicals (3-morpholinosydnonimine), an inhibitor of glutathione synthesis (L-buthionine-sulfoximine) or glutathione reductase (1,3-bis(2-chloroethyl)-1-nitrosourea) in the presence or absence of FBP (3.5 mM). Neuronal viability was determined using an 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. FBP protected neurons against hypoxia-reoxygenation and oxidative stress under conditions of compromised GSH metabolism. The efficacy of FBP depended on duration of hypoxia and was associated with higher intracellular GSH concentration, an effect partly mediated via increased glutathione reductase activity.

Increasing the Glutathione Content in a Chilling-Sensitive Maize Genotype Using Safeners Increased Protection against Chilling-Induced Injury

With the aim of analyzing their protective function against chilling-induced injury, the pools of glutathione and its precursors, cysteine (Cys) and gamma -glutamyl-Cys, were increased in the chilling-sensitive maize (Zea mays) inbred line Penjalinan using a combination of two herbicide safeners. Compared with the controls, the greatest increase in the pool size of the three thiols was detected in the shoots and roots when both safeners were applied at a concentration of 5 muM. This combination increased the relative protection from chilling from 50% to 75%. It is interesting that this increase in the total glutathione (TG) level was accompanied by a rise in glutathione reductase (GR; EC 1.6.4.2) activity. When the most effective safener combination was applied simultaneously with increasing concentrations of buthionine sulfoximine, a specific inhibitor of glutathione synthesis, the total gamma -glutamyl-Cys and TG contents and GR activity were decreased to very low levels and relative protection was lowered from 75% to 44%. During chilling, the ratio of reduced to oxidized thiols first decreased independently of the treatments, but increased again to the initial value in safener-treated seedlings after 7 d at 5 degreesC. Taking all results together resulted in a linear relationship between TG and GR and a biphasic relationship between relative protection and GR or TG, thus demonstrating the relevance of the glutathione levels in protecting maize against chilling-induced injury.

Role of the Ascorbate-Glutathione Cycle of Mitochondria and Peroxisomes in the Senescence of Pea Leaves1

We investigated the relationship between H2O2 metabolism and the senescence process using soluble fractions, mitochondria, and peroxisomes from senescent pea (Pisum sativum L.) leaves. After 11 d of senescence the activities of Mn-superoxide dismutase, dehydroascorbate reductase (DHAR), and glutathione reductase (GR) present in the matrix, and ascorbate peroxidase (APX) and monodehydroascorbate reductase (MDHAR) activities localized in the mitochondrial membrane, were all substantially decreased in mitochondria. The mitochondrial ascorbate and dehydroascorbate pools were reduced, whereas the oxidized glutathione levels were maintained. In senescent leaves the H2O2 content in isolated mitochondria and the NADH- and succinate-dependent production of superoxide (O2·−) radicals by submitochondrial particles increased significantly. However, in peroxisomes from senescent leaves both membrane-bound APX and MDHAR activities were reduced. In the matrix the DHAR activity was enhanced and the GR activity remained unchanged. As a result of senescence, the reduced and the oxidized glutathione pools were considerably increased in peroxisomes. A large increase in the glutathione pool and DHAR activity were also found in soluble fractions of senescent pea leaves, together with a decrease in GR, APX, and MDHAR activities. The differential response to senescence of the mitochondrial and peroxisomal ascorbate-glutathione cycle suggests that mitochondria could be affected by oxidative damage earlier than peroxisomes, which may participate in the cellular oxidative mechanism of leaf senescence longer than mitochondria.

A comparison of the effects of ocular preservatives on mammalian and microbial ATP and glutathione levels

The aim of this study was to investigate the mechanism of action of the preservative sodium chlorite (NaClO2), and the relationship with intracellular glutathione depletion. A detailed comparison of the dose responses of two cultured ocular epithelial cell types and four species of microorganism was carried out, and comparisons were also made with the quaternary ammonium compound benzalkonium chloride (BAK), and the oxidant hydrogen peroxide (H2O2). The viability of mammalian and microbial cells was assessed in the same way, by the measurement of intracellular ATP using a bioluminescence method. Intracellular total glutathione was measured by reaction with 5,5'-dithiobis-2-nitrobenzoic acid in a glutathione reductase-dependent recycling assay. BAK and H2O2 caused complete toxicity to conjunctival and corneal epithelial cells at similar to25 ppm, in contrast to NaClO2 , where >100 ppm was required. The fungi Candida albicans and Alternaria alternata had a higher resistance to NaClO2 than the bacteria Staphyloccus aureus and Pseudomonas aeruginosa , but the bacteria were extremely resistant to H2O2 NaClO2 caused substantial depletion of intracellular glutathione in all cell types, at concentrations ranging from <10 ppm in Pseudomonas , 25-100 ppm in epithelial cells, to >500 ppm in fungal cells. The mechanisms of cytotoxicity of NaClO2 , H2O2 and BAK all appeared to differ. NaClO2 was found to have the best balance of high antibacterial toxicity with low ocular toxicity. The lower toxicity of NaClO2 to the ocular cells, compared with BAK and H2O2 , is in agreement with fewer reported adverse effects of application in the eye.

Dimethylsulfoxide oxidizes glutathione in vitro and in human erythrocytes:kinetic analysis by 1H NMR

The interaction of dimethylsulfoxide (Me2SO) with glutathione was investigated under non-equilibrium conditions in solution using 1H NMR and in intact erythrocytes using 1H spin-echo NMR. In solution the reaction was observed to follow second-order kinetics (Rate = k1[glutathione][Me2SO]) at 300 K pH 7.4, ksol = 4.7 × 10-5 mol -1 L1 s-1. In intact erythrocytes the rate constant for the cellular environment, kcell, was found to be slightly larger at 8.1 × 10-5 mol-1 L1 s-1. Furthermore, the reaction of Me2SO with erythrocyte glutathione showed a biphasic dependence on the Me2SO concentration, with little oxidation of glutathione occurring until the Me2SO concentration exceeded 0.5 mol L-1. The results suggest that at lower concentrations, Me2SO can be effectively removed, most probably by reaction with glutathione, which is regenerated by glutathione reductase, although preferential reaction with other cellular components (e.g., membrane or cellular thiols) cannot be ruled out. Thus the concentrations of Me2SO that are commonly used in cryopreservation of mammalian cells (∼1.4 mol L-1) can cause oxidation of intracellular glutathione.

Glutathione adducts induced by ischemia and deletion of glutaredoxin-1 stabilize HIF-1α and improve limb revascularization

Reactive oxygen species (ROS) are increased in ischemic tissues and necessary for revascularization; however, the mechanism remains unclear. Exposure of cysteine residues to ROS in the presence of glutathione (GSH) generates GSH-protein adducts that are specifically reversed by the cytosolic thioltransferase, glutaredoxin-1 (Glrx). Here, we show that a key angiogenic transcriptional factor hypoxia-inducible factor (HIF)-1α is stabilized by GSH adducts, and the genetic deletion of Glrx improves ischemic revascularization. In mouse muscle C2C12 cells, HIF-1α protein levels are increased by increasing GSH adducts with cell-permeable oxidized GSH (GSSG-ethyl ester) or 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanyl thiocarbonylamino) phenylthiocarbamoylsulfanyl] propionic acid (2-AAPA), an inhibitor of glutathione reductase. A biotin switch assay shows that GSSG-ester-induced HIF-1α contains reversibly modified thiols, and MS confirms GSH adducts on Cys520 (mouse Cys533). In addition, an HIF-1α Cys520 serine mutant is resistant to 2-AAPA–induced HIF-1α stabilization. Furthermore, Glrx overexpression prevents HIF-1α stabilization, whereas Glrx ablation by siRNA increases HIF-1α protein and expression of downstream angiogenic genes. Blood flow recovery after femoral artery ligation is significantly improved in Glrx KO mice, associated with increased levels of GSH-protein adducts, capillary density, vascular endothelial growth factor (VEGF)-A, and HIF-1α in the ischemic muscles. Therefore, Glrx ablation stabilizes HIF-1α by increasing GSH adducts on Cys520 promoting in vivo HIF-1α stabilization, VEGF-A production, and revascularization in the ischemic muscles

Sphingolipid distribution changes with age in the human lens

The formation of an internal barrier to the diffusion of small molecules in the lens during middle age is hypothesized to be a key event in the development of age-related nuclear (ARN) cataract. Changes in membrane lipids with age may be responsible. In this study, we investigated the effect of age on the distribution of sphingomyelins, the most abundant lens phospholipids. Human lens sections were initially analyzed by MALDI mass spectrometry imaging. A distinct annular distribution of the dihydrosphingomyelin, DHSM (d18:0/16:0), in the barrier region was observed in 64- and 70-year-old lenses but not in a 23-year-old lens. An increase in the dihydroceramide, DHCer (d18:0/16:0), in the lens nucleus was also observed in the older lenses. These findings were supported by ESI mass spectrometry analysis of lipid extracts from lenses dissected into outer, barrier, and nuclear regions. A subsequent analysis of 18 lenses ages 20-72 years revealed that sphingomyelin levels increased with age in the barrier region until reaching a plateau at approximately 40 years of age. Such changes in lipid composition will have a significant impact on the physical properties of the fiber cell membranes and may be associated with the formation of a barrier.-Deeley, J. M., J. A. Hankin, M. G. Friedrich, R. C. Murphy, R. J. W. Truscott, T. W. Mitchell, and S. J. Blanksby. Sphingolipid distribution changes with age in the human lens. J. Lipid Res. 2010. 51: 2753-2760.

Regulation of oxidative-stress responsive genes by arecoline in human keratinocytes

Background and Objective: Arecoline, an arecanut alkaloid present in the saliva of betel quid chewers, has been implicated in the pathogenesis of a variety of inflammatory oral diseases, including oral submucous fibrosis and periodontitis. To understand the molecular b asis of arecoline action in epithelial changes associated with these diseases, we investigated the effects of arecoline on human keratinocytes with respect to cell growth regulation and the expression of stress-responsive genes.Material and Methods:Human keratinocyte cells (of the HaCaT cell line) were treated with arecoline, following which cell viability was assessed using the Trypan Blue dye-exclusion assay, cell growth and proliferation were analyzed using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) and 5-bromo-2-deoxyuridine incorporation assays, cell cycle arrest and generation of reactive oxygen species were examined using flow cytometry, and gene expression changes were investigated using the reverse transcription-polymerase chain reaction technique. The role of oxidative stress, muscarinic acetylcholine receptor and mitogen-activated protein kinase (MAPK) pathways were studied using specific inhibitors. Western blot analysis was performed to study p38 MAPK activation.Results:Arecoline induced the generation of reactive oxygen species and cell cycle arrest at the G1/G0 phase in HaCaT cells without affecting the expression of p21/Cip1. Arecoline-induced epithelial cell death at higher concentrations was caused by oxidative trauma without eliciting apoptosis. Sublethal concentrations of arecoline upregulated the expression of the following stress-responsive genes: heme oxygenase-1; ferritin light chain; glucose-6-phosphate dehydrogenase; glutamate-cysteine ligase catalytic subunit; and glutathione reductase.Additionally, there was a dose-dependent induction of interleukin-1alfa mRNA by arecoline via oxidative stress and p38 MAPK activation. Conclusion:our data highlight the role of oxidative stress in arecoline-mediated cell death, gene regulation and inflammatory processes in human keratinocytes.

Neuroprotective activity of Wedelia calendulacea on cerebral ischemia/reperfusion induced oxidative stress in rats

Objective: This study was undertaken to evaluate the neuroprotective activity of Wedelia calendulacea against cerebral ischemia/reperfusion induced oxidative stress in the rats. Materials and Methods: The global cerebral ischemia was induced in male albino Wistar rats by occluding the bilateral carotid arteries for 30 min followed by 1 h and 4 h reperfusion. At various times of reperfusion, the histopathological changes and the levels of malondialdehyde (MDA), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione-s-transferase (GST), and hydrogen peroxide (H(2)O(2)) activity and brain water content were measured. Results: The ischemic changes were preceded by increase in concentration of MDA, hydrogen peroxide and followed by decreased GPx, GR, and GST activity. Treatment with W. calendulacea significantly attenuated ischemia-induced oxidative stress. W. calendulacea administration markedly reversed and restored to near normal level in the groups pre-treated with methanolic extract (250 and 500 mg/kg, given orally in single and double dose/day for 10 days) in dose-dependent way. Similarly, W. calendulacea reversed the brain water content in the ischemia reperfusion animals. The neurodegenaration also conformed by the histopathological changes in the cerebral-ischemic animals. Conclusion: The findings from the present investigation reveal that W. calendulacea protects neurons from global cerebral-ischemic injury in rat by attenuating oxidative stress.

Pharmacological Blockade of Cannabinoid CB1 Receptors in Diet-Induced Obesity Regulates Mitochondrial Dihydrolipoamide Dehydrogenase in Muscle

Cannabinoid CB1 receptors peripherally modulate energy metabolism. Here, we investigated the role of CB1 receptors in the expression of glucose/pyruvate/tricarboxylic acid (TCA) metabolism in rat abdominal muscle. Dihydrolipoamide dehydrogenase (DLD), a flavoprotein component (E3) of alpha-ketoacid dehydrogenase complexes with diaphorase activity in mitochondria, was specifically analyzed. After assessing the effectiveness of the CB1 receptor antagonist AM251 (3 mg kg(-1), 14 days) on food intake and body weight, we could identified seven key enzymes from either glycolytic pathway or TCA cycle-regulated by both diet and CB1 receptor activity-through comprehensive proteomic approaches involving two-dimensional electrophoresis and MALDI-TOF/LC-ESI trap mass spectrometry. These enzymes were glucose 6-phosphate isomerase (GPI), triosephosphate isomerase (TPI), enolase (Eno3), lactate dehydrogenase (LDHa), glyoxalase-1 (Glo1) and the mitochondrial DLD, whose expressions were modified by AM251 in hypercaloric diet-induced obesity. Specifically, AM251 blocked high-carbohydrate diet (HCD)-induced expression of GPI, TPI, Eno3 and LDHa, suggesting a down-regulation of glucose/pyruvate/lactate pathways under glucose availability. AM251 reversed the HCD-inhibited expression of Glo1 and DLD in the muscle, and the DLD and CB1 receptor expression in the mitochondrial fraction. Interestingly, we identified the presence of CB1 receptors at the membrane of striate muscle mitochondria. DLD over-expression was confirmed in muscle of CB1-/- mice. AM251 increased the pyruvate dehydrogenase and glutathione reductase activity in C2C12 myotubes, and the diaphorase/oxidative activity in the mitochondria fraction. These results indicated an up-regulation of methylglyoxal and TCA cycle activity. Findings suggest that CB1 receptors in muscle modulate glucose/pyruvate/lactate pathways and mitochondrial oxidative activity by targeting DLD.

植物体内谷胱甘肽还原酶的功能研究

谷胱甘肽还原酶（GR，EC1.6.4.2）是一重要的抗氧化酶，许多生理学和遗传工程研究都证明GR酶在抗氧化中的重要作用。但改变GR酶怎样影响植物的抗氧化系统却不清楚。GR是抗坏血酸－谷胱甘肽循环途径中的重要组成部分，其功能必然与其密切相关。本文用RNAi技术获得具有较低GR酶活性的转基因烟草，系统测定了非胁迫条件和胁迫条件下抗坏血酸－谷胱甘肽循环的变化，得出以下主要结果： 1．选择一烟草叶绿体GR酶编码基因（X76293, gi: 431954）进行RNAi载体构建，构建好的双元载体转化根癌农杆菌LBA4404，然后侵染转化烟草叶圆片。获得的转基因烟草具有30－70％的GR酶活性。分子检测结果表明GR在RNA和蛋白水平上与GR酶活性的变化一致。文中我们第一次用2-D电泳对烟草中GR同工酶进行分析，并确定发生抑制的GR同工酶在细胞中的定位。2-D电泳后的Western杂交检测到烟草的10种GR同工酶，pI值分布在4.5-6.3，其中3种GR同工酶定位在叶绿体内，其蛋白量占据所有GR酶含量的大部分。RNAi发生在叶绿体内和叶绿体外，表明发生抑制的GR同工酶的基因序列具有很高的同源性。igr转基因烟草在表型上与野生型对照烟草无明显差异。 2．所有igr转基因植株和对照植株中的活性氧（O2-和H2O2）、MDA含量和光合作用都无明显差异，表明正常生长条件下GR酶活性的降低不会引起氧化胁迫。测定正常生长条件下igr转基因烟草中谷胱甘肽库的变化。结果表明与对照烟草相比，GR酶活性降低70％会引起转基因植株中GSH/GSSG比率明显降低，而GSH和GSSG的含量稍有增加；测定抗坏血酸－谷胱甘肽循环的变化，结果显示igr转基因烟草中DHAR和MDHAR的酶活性升高，表明非胁迫条件下较低的GR酶活性可能会诱导抗坏血酸－谷胱甘肽循环不能正常的运转。这一作用可能与改变的谷胱甘肽库有关。GR酶活性降低30％的转基因烟草中未检测到这些变化，表明70％的GR酶活对于非胁迫条件下igr转基因烟草可能是足够的。 3. MV处理结果显示，igr转基因烟草的离体叶圆片和活体植株在MV处理后都发生比对照烟草严重的光漂白作用。igr转基因烟草的活性氧和MDA含量明显高于对照烟草，igr转基因烟草的光合作用明显低于对照烟草。以上这些指标表明igr转基因烟草对MV处理更为敏感。MV处理条件下igr转基因烟草谷胱甘肽的含量明显高于对照烟草，但是GSH/GSSG的比率明显低于对照烟草，GR酶活性仍明显低于对照烟草，表明在MV胁迫条件下igr转基因烟草中较低的GR酶活性不能有效的将GSSG还原生成GSH。igr转基因烟草中较高的谷胱甘肽净含量说明其谷胱甘肽的合成能力提高，但这仍不能补偿胁迫条件下较低GR酶引起的GSH/GSSG比率降低。MV处理条件下igr转基因烟草和对照烟草相比ASC的含量大大降低，导致DHA/ASC明显升高。测定MDHAR和DHAR的结果表明，MV处理后igr转基因烟草的MDHAR酶活性明显降低，这表明较低的GR酶活性引起ASC再生循环受到抑制。MV处理后较低的GR酶还引起igr转基因烟草中APX的活性大大降低。以上这些结果表明MV处理条件下降低GR酶活性会削弱抗坏血酸－谷胱甘肽循环，从而引起活性氧的大量积累，造成严重的氧化伤害。 4．低温处理的结果和MV处理的结果稍有不同。在GR酶活性较高的i2转基因烟草中所有检测指标与对照烟草无明显差异。而GR酶活性较低的i21、i28和i42植株与对照烟草相比表现出明显差异。低温下生长的对照烟草叶绿素含量明显高于i21、i28和i42植株。i21、i28和i42中活性氧（O2-和H2O2）和MDA的含量都明显高于对照烟草，表明低温处理下i21、i28和i42受到更严重的胁迫伤害。与MV处理后的变化相似，低温处理后i21、i28和i42中较低的 GR酶活性导致GSH/GSSG大大降低，ASC再生循环受抑制，APX活性明显降低，从而使抗坏血酸－谷胱甘肽循环不能高效的清除活性氧，导致ROS和MDA的大量积累，造成严重的低温伤害。

水杨酸在水稻抗冷性中的作用

近年来大量研究表明水杨酸（salicylic acid, SA）在植物抵抗生物胁迫与非生物胁迫中都发挥着重要作用。然而在一些单子叶植物如水稻中SA的作用迄今仍不是很清楚。为了更深入地了解SA在水稻抵御冷胁迫中的作用，本研究选用两个抗冷性不同的水稻品种：‘长白九’（Oryza sativa cv. ‘Changbaijiu’）和‘中鉴’（Oryza sativa cv. ‘Zhongjian’）作为实验材料，其中‘长白九’为抗冷性较强的品种，而‘中鉴’为冷敏感的品种。在水稻幼苗长至三叶期后，分别对其施以三种浓度（0.5 mM, 1.0 mM, 2.0 mM）的SA溶液预处理24 h，然后置于5 °C下进行冷处理24 h。形态学观察及各项指标的测定结果表明： 一、冷处理后，‘长白九’和‘中鉴’根与叶片中的SA含量都大幅提高，且结合态SA升高的幅度明显大于其自由态形式。 二、外施不同浓度的SA溶液于水稻根部，24 h后，大量SA尤其是结合态SA积累于根中，且其积累量与处理浓度成正相关;而叶片中积累的SA则较少。 三、形态学及生理指标的测定结果显示，SA预处理没有提高甚至降低了两个水稻品种幼苗的抗冷性。并且SA处理浓度越大，幼苗受到冷伤害程度的越高。 四、对水稻幼苗叶片与根中的抗氧化酶活性进行分析发现，常温下SA处理显著提高了‘长白九’和‘中鉴’根中过氧化氢酶（catalase, CAT）和谷胱甘肽还原酶（glutathione reductase, GR）的活性;而在低温下SA预处理反而降低了两种水稻叶片与根中部分抗氧化酶的活性，推测低温下抗氧化酶活性的下降可能与水稻幼苗抗冷性的降低有关。 五、尽管两个水稻品种具有不同的冷敏感性，然而外施水杨酸均加剧了其低温伤害。分析认为，外施水杨酸后，水稻根部大幅升高的内源SA水平可能加剧了活性氧的产生，破坏了植物细胞内部的氧化还原平衡，从而导致水稻幼苗受到的冷害加重。

Oxidative stress response after prolonged exposure of domestic rabbit to a lower dosage of extracted microcystins

Oxidative stress response after prolonged exposure to a low dose of microcystins (MCs) was studied in liver, kidney and brain of domestic rabbits. Rabbits were treated with extracted MCs (mainly MC-LR and MC-RR) at a dose of 2 MC-LReq. mu g/kg body weight or saline solution every 24 h for 7 or 14 days. During the exposure of MCs, increase of lipid peroxidation (LPO) levels were detected in all the organs studied, while antioxidant enzymes responded differently among different organs. The enzyme activities Of Superoxide dismutase (SOD). catalase (CAT) and glutathione reductase (GR) in liver decreased in the MCs treated animals. In brain, there were obvious changes in glutathione peroxidase (GPx) and GR, while only CAT was obviously influenced in kidney. Therefore, daily exposure at a lower dosage of MCs, which mimicked a natural route of MCs. could also induce obvious oxidative stress in diverse organs of domestic rabbits. The oxidative stress induced by MCs in brain was as serious as in liver and kidney, suggesting that brain may also be a target of MCs in mammals. And it seems that animals may have more time to metabolize the toxins or to form an adaptive response to reduce the adverse effects when exposed to the low dose of MCs. (C) 2008 Elsevier B.V. All rights reserved.