| Objective1-bromopropane (1-BP), also named brompropane and n-propyl bromide, is a colorless transparent organic solvent. Due to its volatile, non-flammol/Lable, and a short half-life of ozone depletion in the air, since the 1990s,1-BP has gradually become the alternative of ozone depleting substances such as CFCS.1-BP has been widely applied in medicine, pesticides, fragrances and other chemical intermediates manufacturing, and it is often used as a fat, resin and paraffin solvents, metal cleaning agent and some electronic instruments. The application and production of 1-BP kept rising year by year in China, the annual output reached 20000 tons in 2008, of which about two-fifths for export. Thus, the population exposed to 1-BP is accordingly expanding.Accumulating 1-BP occupational poisoning patients, epidemiological surveys and experimental researches have demonstrated that 1-BP exposue exerted negative effects both on the central nervous system and peripheral nervous system, with the CNS damage prior to PNS. However, the mechanism of neurotoxicity of 1-BP remains unclear and there was no effective therapeutics for it till now.Studies had shown that 1-BP could significantly disturb the intracellular redox balance due to depletion of GSH in the brain. In vitro culture of macrophage, it was observed that 1-BP could activate the NF-κB. However, whether 1-BP could activate the nuclear factor kappa B (NF-κB) in glia of CNS in vivo and whether NF-κB activation was related to the neurotoxicity of 1-BP, it had not been reported yet. In present study, the contents of GSH, the pathomorphological changes of the brain in rats treated with 1-BP were observed. The activation of NF-κB and Nrf-2, the following events as well were also detected. Meanwhile, the inhibitor of NF-κB-pyrrolidine dithiocarbamate (PDTC) was co-treated with the 1-BP to rat, the neurobehavioral performances, the GSH levels in brains, and the status of neuroinflammol/Lation were analyzed. By comparing the differences between the rats exposed to 1-BP with or without PDTC treatment, we speculated the role of NF-κB in the neurotoxicity of 1-BP.MethodsAfter acclimation for 5 days,60 Male wistar rats (220-240g) were randomly divided into four groups (n= 15 rats/group), including control group,1-BP group, PDTC plus 1-BP group, and PDTC group. The rats in 1-BP and PDTC plus 1-BP group received 800 mg/kg·bw 1-BP (dissolved in corn oil)by gavage respectively, while rats in the control and the PDTC groups received equivalent volume of corn oil. PDTC was given to the rats in PDTC and PDTC plusl-BP group via intraperitoneal injection at a dose of 100mg/kg·bw. Animals in the other groups received volume-equivalent injections of sterile saline on the same schedule.1-BP and PDTC were administered daily, for 13 days at the same time each day. From the 8th day to 13th day of administration,10 rats were randomly taken to Morris water maze (MWM) test in each group. The swimmol/Ling distance and escape latency were used to evaluate the learning ability, the numbers of crossing platform was for memory ability of rats.At the following morning after MWM, the animals were sacrificed. The cerebral cortex were quickly removed, frozen in liquid nitrogen, and kept at -80℃ for western blotting, quantitative real-time PCR (qRT-PCR) and biochemical analysis.The rest of the rats were transcardially perfused with saline followed by fixatives, then, the brains of rats were post-fixated and sliced for immol/Lunofluorescent staining.ResultsThe Morris water maze test showed that the swimmol/Ling distance and escape latency of the rats in 1-BP group was significant greater than that of the control values (P<0.05). Rats of 1-BP groups made significantly fewer platform crossings than rats of the control group (P<0.05). In PDTC plus 1-BP group, the swimmol/Ling distance and escape latency was significant lessen than that of the 1-BP group (.P<0.05). In addition, the PDTC plus 1-BP group rats made more platform crossings than the 1-BP group rats (P<0.05). These results indicated that the impairments in spatial learning and memory induced by the 1-BP could be reversed by PDTC.Compared with the control rats, the glutathione (GSH) levels was markedly decreased in the rats of 1-BP group, being accompanied with the increase of oxidized GSH (GSSG) level, the GSH/GSSG ratio was proportionately decreased (P<0.05). However, there were significantly increasing in the GSH level and GSH/GSSG ratio at PDTC plus 1-BP group compared with 1-BP group, and dramatically decrease in the GSSG level (P<0.05). These data indicated that 1-BP could disturb the redox homeostasis of the brain. The results also demonstrate that the NO levels and iNOS activities were markedly increased in the rats of 1-BP group (P<0.05). Compared with 1-BP group, the nitric monoxide (NO) contents and inducible nitric oxide synthase (iNOS) activities were markedly decreased in the rats of PDTC plus 1-BP group (P<0.05).The results showed that the 1-BP could trigger the NF-κB activation, which were evidenced by the incrase of the level in cytoplasm and the nuclear translocation of NF-κB. The content of inflammol/Latory cytokines such as tumor necrosis factor-a (TNF-a) and interleukin-1β (IL-1β) were also markedly increased in the rats brains of 1-BP group (P<0.05). The NF-κB activation was suppressed by the PDTC treatment, companied with the decrease of the mRNA level of TNF-a and IL-1β.The ctivation of transcription factor NF-E2-related factor-2 (Nrf-2) was also observed both in brain of rates treated with 1-BP and PDTC. The protein levels of the GR and y-glutamate cysteine ligase (GCL) in 1-BP and 1-BP plus PDTC were significantly higher than that of the control group (P<0.05). Contrarily, the activities of the glutathione reductase (GR) and y-GCL in 1-BP group were markly inhibited when compared to control. While, in PDTC plus 1-BP group, activities of the GR and y-GCL was dramatically increase than that of the 1-BP group (P<0.05). The following analysis revealed that the 4-hydroxy-2-nonenal (4-HNE) and malondiadehyde (MDA) modified proteins were significantly increase in 1-BP group. The co-treatemtne of PDTC and 1-BP effectively decreased the oxidative modification of proteins.The immol/Lunofluorescent staining demonstrated that the 1-BP treatment resulted in the activation of microglia and astrocyte in the prefrontal cortex of rats, along with the significant neuronal loss. Following PDTC administration, the reactive glia and neuronal loss was dramatically reversed. These results indicated that the suppression of neuroinflammol/Lation could ameliorate the 1-BP neurotoxicity.Conclusion1.1-BP could activate the astrocyte and microglia, neuronal loss in the prefrontal cortex of rats, final resulted in damages of the learning and memory abilities.2.1-BP could trigger the NF-κB activation, inhibition of NF-κB activation by PDTC could effective reverse the neuroinflammol/Lation and improve the neurobeheavioral performance of rats treated with 1-BP.3.1-BP could activate the Nrf-2 cell signal transduction, which might be one of the compensatory mechanisms of the brain. |
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