Inducible nitric oxide synthase inhibitor, 1400W, mitigates DFP-induced long-term neurotoxicity in the rat model.,Neurobiology of Disease

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Inducible nitric oxide synthase inhibitor, 1400W, mitigates DFP-induced long-term neurotoxicity in the rat model.
Neurobiology of Disease ( IF 5.1 ) Pub Date : 2019-03-30 , DOI: 10.1016/j.nbd.2019.03.031
Marson Putra ₁ , Shaunik Sharma ₁ , Meghan Gage ₁ , Grace Gasser ₂ , Andy Hinojo-Perez ₁ , Ashley Olson ₁ , Adriana Gregory-Flores ₁ , Sreekanth Puttachary ₃ , Chong Wang ₄ , Vellareddy Anantharam ₂ , Thimmasettappa Thippeswamy ₁

Affiliation

Chemical nerve agents (CNA) are increasingly becoming a threat to both civilians and military personnel. CNA-induced acute effects on the nervous system have been known for some time and the long-term consequences are beginning to emerge. In this study, we used diisopropylfluorophosphate (DFP), a seizurogenic CNA to investigate the long-term impact of its acute exposure on the brain and its mitigation by an inducible nitric oxide synthase (iNOS) inhibitor, 1400W as a neuroprotectant in the rat model. Several experimental studies have demonstrated that DFP-induced seizures and/or status epilepticus (SE) causes permanent brain injury, even after the countermeasure medication (atropine, oxime, and diazepam). In the present study, DFP-induced SE caused a significant increase in iNOS and 3-nitrotyrosine (3-NT) at 24 h, 48 h, 7d, and persisted for a long-term (12 weeks post-exposure), which led to the hypothesis that iNOS is a potential therapeutic target in DFP-induced brain injury. To test the hypothesis, we administered 1400W (20 mg/kg, i.m.) or the vehicle twice daily for the first three days of post-exposure. 1400W significantly reduced DFP-induced iNOS and 3-NT upregulation in the hippocampus and piriform cortex, and the serum nitrite levels at 24 h post-exposure. 1400W also prevented DFP-induced mortality in <24 h. The brain immunohistochemistry (IHC) at 7d post-exposure revealed a significant reduction in gliosis and neurodegeneration (NeuN+ FJB positive cells) in the 1400W-treated group. 1400W, in contrast to the vehicle, caused a significant reduction in the epileptiform spiking and spontaneous recurrent seizures (SRS) during 12 weeks of continuous video-EEG study. IHC of brain sections from the same animals revealed a significant reduction in reactive gliosis (both microgliosis and astrogliosis) and neurodegeneration across various brain regions in the 1400W-treated group when compared to the vehicle-treated group. A multiplex assay from hippocampal lysates at 6 weeks post-exposure showed a significant increase in several key pro-inflammatory cytokines/chemokines such as IL-1α, TNFα, IL-1β, IL-2, IL-6, IL-12, IL-17a, MCP-1, LIX, and Eotaxin, and a growth factor, VEGF in the vehicle-treated animals. 1400W significantly suppressed IL-1α, TNFα, IL-2, IL-12, and MCP-1 levels. It also suppressed DFP-induced serum nitrite levels at 6 weeks post-exposure. In the Morris water maze, the vehicle-treated animals spent significantly less time in the target quadrant in a probe trial at 9d post-exposure compared to their time spent in the same quadrant 11 days previously (i.e., 2 days prior to DFP exposure). Such a difference was not observed in the 1400W and control groups. However, learning and short-term memory were unaffected when tested at 10-16d and 28-34d post-exposure. Accelerated rotarod, horizontal bar test, and the forced swim test revealed no significant changes between groups. Overall, the findings from this study suggest that 1400W may be considered as a potential therapeutic agent as a follow-on therapy for CNA exposure, after controlling the acute symptoms, to prevent mortality and some of the long-term neurotoxicity parameters such as epileptiform spiking, SRS, neurodegeneration, reactive gliosis in some brain regions, and certain key proinflammatory cytokines and chemokine.

中文翻译：

诱导型一氧化氮合酶抑制剂1400W在大鼠模型中减轻了DFP诱导的长期神经毒性。

化学神经毒剂（CNA）日益成为对平民和军事人员的威胁。CNA对神经系统的急性影响已为人所知，长期影响已开始显现。在这项研究中，我们使用了二异丙基氟磷酸酯（DFP）（一种能引起精神分裂的CNA）来研究其急性暴露对大脑的长期影响以及诱导型一氧化氮合酶（iNOS）抑制剂1400W作为大鼠模型中的神经保护剂的缓解作用。。几项实验研究表明，即使在采取对策药物（阿托品，肟和地西epa）后，DFP诱发的癫痫发作和/或癫痫持续状态（SE）也会造成永久性脑损伤。在本研究中，DFP诱导的SE在24小时，48小时，7天时引起iNOS和3-硝基酪氨酸（3-NT）的显着增加，并持续了很长时间（暴露后12周），这导致了iNOS是DFP诱导的脑损伤的潜在治疗靶点的假设。为了验证该假设，我们在暴露后的前三天每天两次施用1400W（20 mg / kg，即时）或媒介物。1400W显着降低DFP诱导的海马和梨状皮层iNOS和3-NT上调，以及暴露后24 h的血清亚硝酸盐水平。1400W还可以防止DFP在24小时内导致的死亡率。暴露后7d的大脑免疫组织化学（IHC）显示，在1400W治疗组中，神经胶质增生和神经退行性变（NeuN + FJB阳性细胞）显着减少。1400W，与车辆相比，在连续的12周视频EEG研究中，导致癫痫样发作和自发性复发性癫痫发作（SRS）显着减少。与媒介物治疗组相比，来自相同动物的大脑切片的IHC显示，在1400W治疗组中，各个大脑区域的反应性神经胶质增生（小胶质细胞增生和星形胶质增生）和神经变性显着减少。暴露后6周海马裂解液的多重分析显示，几种关键促炎细胞因子/趋化因子（例如IL-1α，TNFα，IL-1β，IL-2，IL-6，IL-12，IL-12）显着增加媒介物处理的动物中的-17a，MCP-1，LIX和Eotaxin以及生长因子VEGF。1400W可显着抑制IL-1α，TNFα，IL-2，IL-12和MCP-1的水平。在暴露后6周，它也抑制了DFP诱导的血清亚硝酸盐水平。在莫里斯水迷宫中，与暴露前9天（即DFP暴露前2天）在同一象限中所花费的时间相比，在暴露后9天进行探针试验的媒介物处理动物在目标象限中所花费的时间明显减少。。在1400W和对照组中未观察到这种差异。但是，在暴露后10-16d和28-34d进行测试时，学习和短期记忆不会受到影响。加速旋转脚架，单杠测试和强迫游泳测试显示两组之间没有显着变化。总体而言，这项研究的结果表明，在控制了急性症状后，可以使用1400W作为CNA暴露的后续治疗方法，以预防死亡和某些长期的神经毒性参数（如癫痫样发作），作为一种潜在的治疗方法。，SRS，神经退行性疾病，

更新日期：2019-03-30

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