Role of copper in LPS-mediated microglial activation

描述（由申请人提供） 小胶质细胞对维持中枢神经系统（CNS）的内环境至关重要。这些特化的驻留细胞的功能是滋养和支持神经元，并作为响应神经元损伤的第一道防线。 响应于神经病理状态，静止的小胶质细胞经历一系列变化，导致释放促炎和细胞毒性介质以去除病原体。在通过吞噬作用和/或去除毒素和毒物清除受损细胞后，小胶质细胞恢复到静息状态或经历程序性细胞死亡。因此，小胶质细胞根据其周围环境表现出不同的表型。适当表型的表达对于成功去除病原体和限制对周围神经元的损伤至关重要。慢性小胶质细胞活化已在多种神经退行性疾病中观察到，但迄今为止，尚不清楚小胶质细胞活化是由于保证其活化状态的持续神经元变性，还是由于小胶质细胞功能障碍，包括未能上调或下调细胞毒性介质（包括一氧化氮（NO））的释放。 NO既是一种有效的细胞毒性介质，也是小胶质细胞内细胞信号传导的关键调节因子。研究者的假设是，小胶质细胞对毒素暴露的表型反应依赖于NO的代谢命运。氧化还原活性过渡金属已被提出作为神经退行性疾病，包括阿尔茨海默氏症，帕金森氏症和肌萎缩侧索硬化症的重要因素。过渡金属铜的水平受到严格管制，偏差将通过改变细胞的氧化还原环境来改变NO信号传导，特别是关于硫醇。研究者建议研究铜改变铜刺激的NO信号传导的机制，从而改变小胶质细胞的表型反应。在Andrew Gow博士实验室的指导阶段，研究人员将研究铜对永生化BV-2和原代小胶质细胞培养物中表型分化的影响。 特别是，她将研究铜如何改变关键信号分子和S-亚硝基化谱，以应对急性毒素挑战，以及铜的存在如何干扰适应性炎症表型的采用。该奖项的独立阶段将建立在辅导阶段获得的调查结果的基础上。在该阶段期间，研究者将研究慢性铜过载对tx j小鼠中响应于全身LPS激发的特定解剖脑结构中的小胶质细胞表型变化的影响。还将研究慢性铜超负荷对小胶质细胞表型的影响是永久性的还是在去除过量铜后可以逆转。 公共卫生相关性：小胶质细胞是大脑中的常驻免疫细胞，在那里它们提供了应对神经损伤的第一道防线。为了应对周围环境的变化，小胶质细胞的激活状态可能会发生变化，也就是说，它们可能会发生表型变化。这些变化通常伴随着促炎或抗炎产物的释放，这将影响神经损伤的结果。在这个研究项目中，研究人员建议定义一氧化氮（激活的小胶质细胞的细胞毒性产物之一）在小胶质细胞表型分化中的作用，以及铜（其水平在各种脑病理中升高）如何干扰蛋白质合成的S-亚硝基化以响应毒素。了解控制小胶质细胞表型变化的条件可能会导致神经退行性疾病的新疗法的发展，其中小胶质细胞处于慢性激活状态。

DESCRIPTION (provided by applicant) Microglia are critical to maintaining the internal environment of the central nervous system (CNS). These specialized resident cells function to nourish and support neurons and to act as a first line of defense in response to neuronal injury. In response to a neuropathological state, quiescent microglia undergo a series of changes that result in the release of pro-inflammatory and cytotoxic mediators for the removal of the pathogen. Upon clearance of injured cells by phagocytosis and/or the removal of toxin and toxicants, microglia return to a resting state or undergo programmed cell death. Microglia, therefore, exhibit different phenotypes depending on their surrounding environment. Expression of the appropriate phenotype is critical to the successful removal of the pathogen and to limiting damage to surrounding neurons. Chronic microglial activation has been observed in a variety of neurodegenerative diseases but, to date, it is not clear whether microglial activation is due to a persistent neuronal degeneration that warrants their activated state, or to microglial dysfunction, including a failure to either up-regulate or down-regulate the release of cytotoxic mediators including nitric oxide (NO). NO is both a potent cytotoxic mediator and a key regulator of cellular signaling within microglia. The investigator's hypothesis is that the phenotypic response of microglia to toxin exposure is dependent on the metabolic fate of NO. Redox active transition metals have been proposed as important factors in neurodegenerative diseases including Alzheimer's, Parkinson's and amiotropic lateral sclerosis. Levels of the transition metal copper are strictly regulated and deviations will alter NO signaling by changing the redox environment of the cell, particularly in reference to thiols. The investigator proposes to investigate the mechanisms by which copper alters copper- stimulated NO signaling and, thus, the phenotypic response of microglia. During the mentored phase of the award in the laboratory of Dr. Andrew Gow, the investigator will investigate the effects of copper on phenotypic differentiation in immortalized BV-2 and in primary microglia cell cultures. In particular, she will examine how copper alters key-signaling molecules and the S-nitrosylation profile in response to an acute toxin challenge and how the presence of copper might interfere with the adoption of an adaptive inflammatory phenotype. The independent phase of the award will build upon the findings obtained during the mentored phase. During this phase the investigator will investigate the effects of chronic copper overload on microglia phenotypic changes in specific anatomical brain structures in response to systemic LPS challenge in the tx j mouse. The effects of chronic copper overload will also be investigated with respect to whether the effects on microglia phenotype are permanent or can be reversed after excess copper has been removed. Public Health Relevance: Microglia are the resident immune cells in the brain where they provide the first line of defense in response to neurological insults. In response to changes in the surrounding environment, the state of activation of microglia may change, that is, they may undergo phenotypic changes. These changes are usually accompanied by the release of pro- or anti-inflammatory products that will affect the outcome of the neurological insult. In this research project the investigators propose to define the role that nitric oxide, one of the cytotoxic products of activated microglia, plays in the phenotypic differentiation of microglia and how copper (whose levels are elevated in a variety of brain pathologies), may interfere with the S-nitrosylation of protein synthesis in response to a toxin. Understanding the conditions that govern phenotypic changes in microglia may lead to the development of novel therapeutics in neurodegenerative diseases where microglia are in a chronic state of activation.