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This study explored the protective effects of endoplasmic reticulum (ER) stress preconditioning induced by 2-deoxy-d-glucose (2-DG) or oxidized dithiothreitol (DTTox) on acrylonitrile (AN)-induced cytotocity in primary rat astrocytes. Cells were pretreated with 2-DG or DTTox for different times at various concentration. Next, astrocytes were treated with 2.5mM AN for an additional 12h. Cell viability and cytotoxicity were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction and lactate dehydrogenase (LDH) leakage, respectively. Reactive oxygen species (ROS) and mitochondrial membrane potential (ΔΨm) were determined. Expression of glucose-regulated protein 78 (GRP78), phosphorylated-eukaryotic translation initiation factor 2α (p-eIF2α), microtubule-associated protein light chain 3 (LC3), P62, and Beclin1 were used to assess autophagy. In addition, 3-methyadenine (3-MA), an autophagy-specific inhibitor, was used to assess the role of autophagy in ER stress preconditioning-induced protection against AN cytotoxicity. The results showed that AN alone significantly decreased astrocytic viability and enhanced cytotoxicity. Compared to the AN-alone group, preconditioning with 2-DG or DTTox significantly increased cell viability and reduced cytotoxicity to indistinguishable levels. Decreased ROS generation and increased ΔΨm were also inherent to ER stress preconditioning with these compounds. Furthermore, autophagy was activated by both 2-DG and DTTox. Blockage of autophagy attenuated the protection afforded by 2-DG or DTTox preconditioning in AN-treated astrocytes. These results establish that ER stress preconditioning affords cellular protection against AN, and that activation of autophagy mediates the cytoprotection. Modulation of ER stress and resultant activation of autophagy may be a novel target for to ameliorate AN toxicity.

Objective This study was designed to examine the differential protection of pre- versus post-treatment with three different antioxidants, curcumin (CUR), Trolox, and N-acetylcysteine (NAC), on acrylonitrile (AN)-induced cytotoxicity in primary rat astrocytes. Methods Primary astrocyte cultures were treated with CUR, Trolox and NAC for 4 h prior to, or following 24 h treatment with AN (2.5 mM). 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), lactate dehydrogenase (LDH) release, lipid peroxidation, glutathione, reactive oxygen species (ROS) and mitochondrial membrane potential were measured to evaluate protection associated with the three antioxidants. Knockdown of Nrf2 expression by liposome transfection with siRNA was used to confirm the role of Nrf2 activation in the protection associated with the three antioxidants. Results Compared with AN treatment alone, pre-treatment with CUR at either concentration significantly increased cell viability and mitochondrial membrane potential, and reduced glutathione levels; lipid peroxidation and ROS production were significantly decreased as well. NAC also showed significant efficacy in attenuating AN-induced toxicity at higher concentration. However, pre-treatment with Trolox failed to ameliorate the AN-induced toxicity. When post-treatment with Trolox, this antioxidant led to significant protective effects at both concentrations, while CUR and NAC were efficacious only at the higher concentrations. Knockdown of Nrf2 only abolished the protective effects of CUR pre-treatment on AN-induced cytotoxicity, while the protective effects of NAC and Trolox pre-treatment groups showed no differences between the Nrf2-knockdown and non-knockdown treatments. Conclusions The selected antioxidants exert differential cellular protection when administered prior or subsequent to AN-induced cytotoxic events in decreasing cellular viability, antioxidative capacity and mitochondrial function, enhanced cytotoxicity and ROS production. These results suggest that antioxidants should be carefully chosen for their efficacy in preventing or diminishing oxidative damage caused by AN. The differential effect of pre- and post-treatment may be attributed to activation of the Nrf2 signaling pathway.

Acrylonitrile (AN) is widely used in the manufacturing of fibers, plastics and pharmaceuticals. Free radical-mediated lipid peroxidation is implicated in the toxicity of AN. The present study was designed to examine the ability of curcumin, a natural polyphenolic compound, to attenuate acute AN-induced lipid peroxidation in the brain and liver of rats. Male Sprague-Dawley rats were orally administered curcumin at doses of 0 (olive oil control), 50 or 100 mg/kg bodyweight daily for 7 consecutive days. Two hours after the last dose of curcumin, rats received an intraperitoneal injection of 50 mg AN/kg bodyweight. Acute exposure to AN significantly increased the generation of lipid peroxidation products, reflected by high levels of malondialdehyde (MDA) both in the brain and liver. These increases were accompanied by a significant decrease in reduced glutathione (GSH) content and a significant reduction in catalase (CAT) activity in the same tissues. No consistent changes in superoxide dismutase (SOD) activity were observed between the control and AN-treatment groups in both tissues. Pretreatment with curcumin reversed the AN-induced effects, reducing the levels of MDA and enhancing CAT activity and increasing reduced GSH content both in the brain and liver. Furthermore, curcumin effectively prevented AN-induced decrease in cytochrome c oxidase activity in both liver and brain. These results establish that curcumin pretreatment has a beneficial role in mitigating AN-induced oxidative stress both in the brains and livers of exposed rats and these effects are mediated independently of cytochrome P450 2E1 inhibition. Accordingly, curcumin should be considered as a potential safe and effective approach in attenuating the adverse effects produced by AN-related toxicants.

Paradichlorobenzene (pDCB) has been used as a space deodorant and moth repellant, as well as an intermediate in the chemical industry. Given its broad applications and high volatility, considerable concern exists regarding the adverse health effects of pDCB in the home and the workplace. In this study, changes in lipid peroxidation, antioxidants, and trace element levels in the liver and kidney of pDCB-treated mice were investigated to determine their roles in toxicity. Mice were orally gavaged once daily for seven consecutive days with pDCB (0 (corn oil control), 450, and 900 mg/kg). The level of malondialdehyde (MDA), an end product of lipid peroxidation, markedly increased in the high-dose pDCB group in both the liver and kidney compared with the control group. Changes in hepatic levels of reduced glutathione (GSH) in the pDCB groups were indistinguishable from the control group, while renal levels of reduced GSH in the high-dose pDCB group were significantly lowered in comparison to the control and the low-dose groups. Superoxide dismutase (SOD) activity in the liver of mice treated with pDCB showed a downward trend, whereas there was no consistent trend associated with changes in SOD activity in the kidney. Additionally, renal iron levels in the high-dose pDCB group were significantly decreased compared with the low-dose group and the controls, whereas hepatic iron content in the low-dose pDCB group was significantly lower compared with the controls. Selenium and zinc levels in the kidney were both significantly decreased in the high-dose pDCB group vs. the control and low-dose groups. There were no treatment-induced changes in copper levels in either the kidney or liver. However, a significant increase was found in the liver zinc/copper ratio in the high-dose pDCB group vs. the controls. In addition, blood zinc levels showed a downward trend with increased pDCB dosage. These results suggest that pDCB toxicity is mediated by oxidative damage and tissue-specific alterations in trace element levels both in the liver and the kidney of mice.

The mechanism of toxicity of acrylonitrile (AN) has not been fully defined. The research described herein was undertaken to investigate the possible effects of AN on the levels of metallic elements in liver and brain of mice. Thirty-two mice were randomly assigned to four separate groups and treated intraperitoneal (i.p.) once daily for 1 week. Mice in the control group received normal saline, and mice in the three exposure groups received 5, 10, or 20 mg AN/kg b.w. Samples of brain and liver were collected immediately after decapitation. Tissue levels of trace elements (zinc, copper, iron) were determined with flame atomic absorption spectrophotometer or double channel atomic fluorescence absorption spectrophotometer (selenium). Mean brain weights of AN-treated mice were increased as a function of dose compared to controls, but there was no significant change in the ratio of liver/body weight in the four groups. While mean brain zinc decreased as a function of AN dosage, mean liver zinc of the low-dose group significantly increased (p0.05); mean liver copper in the medium-dose AN group was significantly higher compared to controls (p0.05); however, mean brain copper was increased, but the difference did not attain statistical significance in the three AN groups when compared with the controls (p0.05). Mean brain iron levels were significantly decreased in the middle-dose AN group (p0.05), but there were no consistent changes in liver iron. Tissue levels of selenium in brain and liver were similar for the control and AN treatment groups. AN induces significant and differential changes in the levels of zinc, copper, and iron in brain and liver. These changes likely play a pivotal role in mediating AN toxicity, most likely via changes in cellular redox status.

This study was carried out on rodents, to explore the neurobehavioral effects of acrylonitrile (AN) administered in drinking water. Thirty, male, Sprague-Dawley rats were randomly divided into three groups: two exposure groups (50 and 200 ppm AN), and one control group (tap water without AN). Three tests, including the open field test, rotarod test and spatial water maze, were applied to evaluate locomotor activities, motor co-ordination and learning and memory, respectively, prior to initiation of the treatment, and at Week 4, 8 and 12 postexposure. There were no consistent changes in the open field test, except for locomotion and grooming episodes. In the rotarod test, AN significantly decreased the latencies to fall in a dose and time-dependent manner. In the spatial water maze test, rats exposed to AN for 12 weeks had significantly more training times and longer escape latencies than control animals. These findings indicate that oral exposure to AN induces neurobehavioral alterations in rats.

Autophagy is an evolutionarily conserved process in which cytoplasmic proteins and organelles are degraded and recycled for reuse. There are numerous reports on the role of autophagy in cell growth and death; however, the role of autophagy in methylmercury (MeHg)-induced neurotoxicity has yet to be identified. We studied the role of autophagy in MeHg-induced neurotoxicity in astrocytes. MeHg reduced astrocytic viability in a concentration- and time-dependent manner, and induced apoptosis. Pharmacological inhibition of autophagy with 3-methyladenine (3-MA) or chloroquine (CQ), as well as the silencing of the autophagy-related protein 5 (Atg5), increased MeHg-induced cytotoxicity and the ratio of apoptotic astrocytes. Conversely, Rapamycin, an autophagy inducer, along with as N-acetyl-L-cysteine (NAC), a precursor of reduced glutathione (GSH), decreased MeHg-induced toxicity and the ratio of apoptotic astrocytes. These results indicated that MeHg-induced neurotoxicity was reduced, at least in part, through the activation of autophagy. Accordingly, modulation of autophagy may offer a new avenue for attenuating MeHg-induced neurotoxicity.

Acrylonitrile(AN) is a neurotoxin both in animals and humans, but its effects on acetylcholinesterase (AChE) activity remain controversial. This study aimed to determine the dose-response effects of AN on AChE activity and the modulatory role of ethanol pretreatment. A total of 144 Kunming mice were randomly divided into 18 groups: nine groups received 5% ethanol in their drinking water, and the remaining nine groups received regular tap water. One week later, both the ethanol and tap water only groups were given an intraperitoneal injection of AN at the following doses: 0 (control), 0.156, 0.3125, 0.625, 1.25, 2.5, 5, 10 or 20 mg AN/kg body weight. AChE activity was determined on whole blood and brain 24 h later. Blood AChE activity was higher in AN-injected mice than in controls at all doses. AChE activity in blood increased in a dose-dependent manner, peaking at 0.156 mg/kg, after which a gradual decrease ensued, displaying a β-typed dose-response relationship. In contrast, brain AChE activity, following a single AN injection, was consistently lower than in control mice, and continued to fall up to a dose of 0.313 mg/kg, and thereafter increased gradually with higher doses. Mice receiving a 20 mg/kg dose of AN exhibited AChE brain activity indistinguishable from that of control mice, demonstrating a typical U-typed dose-response relationship. The activity of AChE in the blood and brain of the AN + ethanol-treated groups displayed a shift to the right, and the magnitude of the decrease in AChE activity induced by AN was attenuated relative to the AN-only group. These results suggest that AN affects AChE activity in both mouse blood and brain in a hormetic manner. Pretreatment with ethanol modifies the effect of AN on AChE, indicating that parent AN has a more prominent role than its metabolites in modulating enzyme activity.

The present study was designed to examine the effects of the inhibition or induction of CYP2E1 activity on acute acrylonitrile (AN) toxicity in rats. Increased or decreased hepatic CYP2E1 activity was achieved by pretreatment with acetone or trans-1,2-dichloroethylene (DCE), respectively. AN (50 mg/kg) was administered by intraperitoneal injection. Onset of convulsions and death were observed in rats with increased CYP2E1 activity, whereas convulsions and death did not appear in rats within 1 h after treatment with AN alone. Convulsions occurred in all AN-treated animals with increased CYP2E1 activity at approximately 18 min. The levels of cyanide (CN(-)), a terminal metabolite of AN, were significantly increased in the brains and livers of the AN-treated rats with increased CYP2E1 activity, compared with the levels in rats treated with AN alone, DCE + AN or acetone + DCE + AN. The cytochrome c oxidase (CcOx) activities in the brains and livers of the rats treated with AN or AN + acetone were significantly lower than those in the normal control rats and the rats treated with DCE, whereas the CcOx activities in the brains and livers of rats with decreased CYP2E1 activity were significantly higher than those in AN-treated rats. Brain lipid peroxidation was enhanced, and the antioxidant capacity was significantly compromised in rats with decreased CYP2E1 activity compared with rats with normal or increased CYP2E1 activity. Therefore, inhibition of CYP2E1 and simultaneous antioxidant therapy should be considered as supplementary therapeutic interventions in acute AN intoxication cases with higher CYP2E1 activity, thus a longer window of opportunity would be got to offer further emergency medication.

While the adverse effects of acrylonitrile (AN) on the central nervous system (CNS) are known to be mediated, at least in part, by the generation of free radicals and oxidative stress, there is a paucity of data on region-specific alterations in biomarkers of oxidative stress in the brain of AN-exposed animals. The present study was designed to examine the effects of AN on biomarkers of oxidative stress in several brain regions of adult Sprague-Dawley rats. Daily intraperitoneal (i.p.) treatment of animals to 0 (control, normal saline solution), 25, 50 or 75mgAN/kg body weight for 7 days resulted in statistically significant (p