| Objective1-Bromopropane (1-BP) is widely used as a substitute for Freon compounds because of not destroying the ozone layer. With the numerous use of 1-BP, more and more workers were exposed to 1-BP, they were absorbed into the body through the skin and respiratory and produced some toxic effects, such as liver toxicity, reproductive toxicity, and nervous system toxicity, nervous system toxicity is the most sensitive, in other words, the most sensitive target in 1-BP toxicity is the central system. But the 1-BP neurotoxicity study is too little, the protective measures and related occupations poisoning treatment measures are not systematic, so it is urgent to develop the drug in the treatment 1-BP neurotoxicity. Organic solvents have a common feature that is soluble in fat, and they have a high affinity for nerve tissue, 1-BP is no exception. The main symptoms of central nervous system toxicity of organic solvents in addition to dizziness, nausea, headache, vomiting, fatigue, difficulty concentrating, memory loss, irritability and so on. But so many nervous system toxicity cannot be reproduced in animals. In the present study, we detected SD learning and memory ability by Morris water maze tests in SD rats, partly reflecting the central nervous system toxicity, and explore the nervous system poisoning prevention and treatment in 1-BP central nervous toxicity.There are two approaches in the treatment of 1-BP toxicity, one is for the cause of treatment, the workers are far away from 1-BP environment as soon as possible, nervous system function can be freely restored in mild 1-BP poisoning. That is poor in severe 1-BP poisoning treatment for the cause of treatment. The other is drug treatment,1-BP poisoning is no specific antidote in the current situation, the way is mainly through symptomatic and supportive treatment, and the prognosis is poor in severe cases. It is unclear in the mechanism of 1-BP poisoning CNS changes and learning and memory function damage, but more and more studies show that 1-BP nervous system toxicity may be caused by oxidative stress.In the study, we investigate four antioxidants in order to find a cure for 1-BP poisoning learning and memory dysfunction. Melatonin can eliminate free radicals, inhibit the lipid peroxidation of cell structure, prevent DNA from damage, decrease the content of peroxides in vivo. The active ingredients of Ginkgo biloba extract (EGb) are flavonoids and lactones, the main components have synergistic antioxidant function, scavenge free radicals, improve cerebral blood circulation, and improve cognitive function. Garlic oil is not limited to food and health care products, there are many medicinal value in the garlic oil including the inhibition of bacteria, fungi, viruses, blood pressure, blood cholesterol, platelet aggregation, free radical scavenging and other effects. Allicin is known as natural spectrum antibiotic, allicin has its unique pharmacological activity and can be anti-cancer, lowering cholesterol, anti-platelet aggregation, lowering blood pressure and blood fat, anti-aging and memory decline. The purpose of the study is to screen the therapeutic drugs of 1-BP toxicity and explore the mechanism to facilitate the clinical use.In this study, we can imitate learning and memory damage in clinical practice due to long-term clinical exposure of 1-BP by the long oral administration of 1-BP. Four different antioxidants are used to improve learning and memory dysfunction with 1-BP central system toxicity. Take Melatonin for example, we study the improvement of learning and memory dysfunction induced by 600mg/kg 1-BP at the different doses of Melatonin by Morris water maze, and explore the mechanisms to improve neurological dysfunction caused by exposure to 1-BP by oxidative stress indexes, HO-1 protein expression, pathological and other indicators. These data can support theoretical foundation for the treatment of 1-BP poisoning.Methods1 The establishment of CNS Neurotoxicity animal model of 1-BP intoxication48 male SD rats were maintained for 5 days and then randomly assigned to 4 groups, including corn oil group,1-BP (200mg/kg bw) group,1-BP (400mg/kg bw) group,1-BP (600mg/kg bw) group. The rats were administered once a day for consecutive 27 days. The dose is calculated by measuring the latest body weight, and the administration volume is 2 mL/kg. The weight of the rats were measured on administration of 1,4,7,10,13,16,19,22,27 days and statistical food consumption per rat was calculated on 1-7,7-14,14-21 days. From Day22-27, Morris water maze test was used to determine the learning and memory in rats.D22 was as the training day and not included in the statistics, Day23-26 was for navigation test, D27 was as spatial navigation test. The remaining two animals per group were anesthetized with the injection of 30% urethane by the intraperitoneal administration, and were intracardially perfused with the saline and 4% paraformaldehyde, then carefully removed the brains into 4% paraformaldehyde. Frozen sections and paraffin sections were done. Hippocampal morphology of rats was observed. This study was evaluated whether learning and memory impairment animal model by behavioral indicators and pathology morphology is successful or not.2 The intervention study of four antioxidants in CNS Neurotoxicity animal model of 1-BP intoxication60 male SD rats were maintained for 5 days and then randomly assigned to 6 groups, including corn oil group,1-bromopropane group (orally 600mg/kg/day), melatonin (intraperitoneal injection of 10mg/kg/day)+1-bromopropane group, Ginkgo biloba extract (orally 100mg/kg/day)+1-bromopropane group,allicin (orally 60mg/kg/day)+1-bromopropane group, garlic oil(orally 60mg/kg/day)+1-bromopropane group. The administration was kept for 27 consecutive days. Administering the test compound was 1 hour before 1-BP administration, wherein melatonin intervention group was generally administrated at 6:00 pm. D22 was as the training day and not included in the statistics, Day23-26 was for navigation test, D27 was as spatial navigation test. In this study, the interference of four antioxidants in learning and memory impairment animal model of 1-BP intoxication was evaluated.3 The protective effects and mechanisms of Melatonin on central nervous system damage caused by 1-Bromopropane60 male SD rats were maintained for 5 days and then randomly assigned to 6 groups, including corn oil group,1-bromopropane group (orally 600mg/kg/day), melatonin control group (intraperitoneal injection of 10mg/kg/day), melatonin (intraperitoneal injection of 2.5mg/kg/day)+1-bromopropane group, melatonin (intraperitoneal injection of 5mg/kg/day)+1-bromopropane group, melatonin (intraperitoneal injection lOmg/kg/day)+1-bromopropane group. The administration was kept for 27 consecutive days. Melatonin intervention group was generally administrated at 6:00 pm. D22 was as the training day and not included in the statistics, Day23-26 was for navigation test, D27 was as spatial navigation test. After the end of the experiment, anesthetized with an excess of sodium pentobarbital, sacrificed, carefully removed the brain and isolated hippocampus, cortex and cerebellum, quick-frozen in liquid nitrogen, transferred to α-80℃ refrigerator. The remaining five animals per group were anesthetized with the injection of 30% urethane by the intraperitoneal administration, and were intracardially perfused with the saline and 4% paraformaldehyde, then carefully removed the brains into 4% paraformaldehyde.The study tested the interference of melatonin on learning and memory impairment in 1-BP intoxication by Morris water maze, then detected hippocampus GSH, GSSG, MDA, H2O2 content and SOD activity by biochemical methods, examined hippocampal neurons changes in CA3 region by immunohistochemistry study, detected the reasons of the neurons loss by TUNEL kit, detected HO-1 expression using western blotting.Results1 The establishment of CNS Neurotoxicity animal model of 1-BP intoxication1.1 the influence of learning and memory in 1-BP intoxication by Morris water maze in SD ratsIn Morris water maze, compared to the corn oil control group, the escape latency and swimming distance was significantly longer and the number of times crossed the target and the time percentage of the target quadrant decreased significantly in 1-BP 200mg/kg group,1-BP 400mg/kg group and 1-BP 600mg/kg group, there are differences between the groups, especially 1-BP high dose (600mg/kg) was the most obvious, the data suggested 1-BP could impair the learning and memory in SD rats. 1.2 Hippocampal neuron morphological changes associated with learning and memory by pathological studyThe hippocampus as the key parts of Learning and memory closely related was examined by Thionine staining and HE staining, and the data showed that 1-BP obviously caused cells in hippocampal CA1 and CA3 area reduction apparently, irregular, Cell vacuoles, deep stained, shrunken nuclei and the cell gap became larger.2 The intervention study of four antioxidants in CNS Neurotoxicity animal model of 1-BP intoxicationIn the navigation test, the daily escape latency and swimming distance substantially gradually shorter with the number of morris water maze days increased in all experimental animals, it is a decreasing trend, and the swimming speed did did not differ significantly in all groups. Melatonin, allicin and garlic oil can reduce the learning impairment induced by 1-BP, while Ginkgo biloba extract is not obvious. Three kinds of antioxidants can protect from spatial learning ability impairment caused by 1-BP, melatonin is the most apparent.In the spatial navigation test, compared with the 1-BP group, melatonin intervention group significantly increased the number of times crossed the target and the time percentage of the target quadrant, allicin and garlic oil intervention group also played a similar effect, while no significant change was noted in ginkgo biloba extract intervention group. These results indicate that melatonin, allicin and garlic oil improve spatial memory damage exposed to 1-BP and the effect is remarkable.Given the results in navigation test and spatial navigation test, melatonin, allicin and garlic oil can improve learning and memory disorders induced by 1-BP, melatonin is the most obvious.3 The protective effects and mechanisms of Melatonin on central nervous system damage caused by 1-Bromopropane3.1 Melatonin improved CNS Neurotoxicity animal model of 1-BP intoxicationMorris water maze test results showed that melatonin can dose-dependently improve learning ability damage induced by 1-BP obtained from the escape latency and swimming distance, the high dose is the most obvious. Melatonin intervention group significantly increased the number of times crossed the target in middle and high dose. Melatonin intervention group significantly increased the time percentage of the target quadrant in high dose. These results showed that melatonin can dose-dependently improve spatial memory disorder caused by 1-BP, and high dose group was significantly apparent.3.2 Melatonin can significantly increase GSH content and the ratio of GSH/GSSG in hippocampal tissue of SD rats.Compared with 1-BP group, Different doses of melatonin can alliviate the loss of GSH content, while the GSH content was increased to as much as 163.6 percent in melatonin high dose group, a statistically significant difference was noted. The high dose of melatonin intervention group significantly increased GSH/GSSG ratio.3.3 Melatonin significantly reduced MDA and H2O2 levels and increased the activity of SOD in the hippocampus of SD rats.Compared with the 1-BP group, different doses of melatonin showed some antioxidant effects, MDA content was significantly reduced by 36% and 44% in melatonin middle and high dose group, respectively, and has a significant difference. Compared with 1-BP exposure group, melatonin dose-dependently decreased hydrogen peroxide levels, and reduced by 21%,22%,36% in low, middle and high dose of melatonin. Although no significant dose-dependent change was observed, there are significant differences in all groups. Melatonin significantly increased the antioxidant enzyme superoxide dismutase activity by 23% and 33% in middle and high dose respectively, and a significant difference was noted.3.4 Hippocampus GSH/GSSG ratio and MDA and cognitive function has a good correlationCorrelation analysis showed that and GSH/GSSG ratio reduction in SD rat hippocampus has a positive correlation, while cognitive impairment and elevated levels of MDA in SD rat hippocampus has a negatively correlation by Spearman correlation.3.5 Melatonin improved the number of neurons decreased by 1-BP in hippocampal CA3 region in SD ratsIn Corn oil control group hippocampal CA3 region, neurons are intact, sharp edges, neat cells, tightly packed cells, more cell layers. In 1-BP exposure hippocampal CA3 region, neurons edge is unclear, the cell gap becomes larger, loosely arranged, a large number of neuron has lost. Melatonin (10mg/kg bw)+1-BP were similar to Corn oil control group hippocampal CA3. The results showed that melatonin can improve the hippocampal CA3 neuron loss induced by 1-BP in SD rats.3.6 Melatonin can improve hippocampal neurons apoptosis induced by 1-BP.There are no stained violet blue cells in neurons, whereas a large number of TUNEL-positive neurons violet blue cells were observed in 1-BP hippocampal neurons. There were no significant differences between corn oil control hippocampal neurons and melatonin (10mg/kg bw)+1-BP group hippocampal neurons.3.7 Melatonin reduced HO-1 content in hippocampusCompared with the 1-BP exposure group, different doses of melatonin dose-dependently decreased HO-1 content in SD rat hippocampus.Conclusion1.1-BP can induce CNS intoxicity in a dose dependent manner in SD rats, which may closely relate to hippocampal CA3 neuron apoptosis.2. Melatonin, allicin and garlic oil can improve the central nervous system damage caused by 1-BP, of which melatonin is the most obvious.3.1-BP can decrease GSH content, GSH/GSSG ratio and SOD levels in SD rat hippocampus,1-BP can increase MDA content and H2O2 levels SD rat hippocampus, while melatonin can antagonize the oxidative stress damage.4.1-BP can increase HO-1 content caused by oxidative stress in the body as a protective defense mechanism, melatonin can reduce the hippocampus of HO-1 content, protecting the body from oxidative shock damage. |
PDF Full Text Request