Elucidation of nitric oxide resistance mechanisms in motor neurons

说明(申请人提供)：一氧化氮(NO)是一种被中枢神经系统用于正常生理功能的自由基。然而，当它在不适当的位置或过量释放时，不会导致硝基酪氨酸的形成，硝基酪氨酸是NO介导的损伤的标志，并导致大量细胞死亡。细胞死亡和硝基酪氨酸形成可见于脊髓损伤和神经退行性疾病，如阿尔茨海默氏症、多发性硬化症(MS)和肌萎缩侧索硬化症(ALS)。有趣的是，用亚毒性剂量的NO预处理的运动神经元细胞对正常毒性剂量的NO获得了显著的抵抗，这种现象我们称之为诱导适应性抵抗(IAR)。此外，当暴露在有毒剂量的NO中时，适应的细胞几乎没有硝基酪氨酸的形成。IAR依赖于血红素代谢酶--血红素加氧酶1(HO-1)。这项资助的总体目标是阐明和剖析IAR现象，并通过这样做，开始了解CNS中的天然抵抗机制，这些机制可以用来保护神经元免受中枢神经系统损伤和神经退行性疾病中出现的NO介导的损伤。目的是：1.确定NO诱导的细胞死亡是否是由于相关的细胞蛋白硝化所致。如果是这样的话，a)阻止3NY的形成也应该阻止NO的杀戮，b)在没有处理的情况下，细胞蛋白质的硝化应该足以杀死细胞。2.确定HO1是如何诱导对NO产生适应性抗性的。HO1可能通过a诱导对NO的适应性抗性。)降低细胞内血红素浓度，b.)通过产生血红素代谢物，如胆红素和一氧化碳，c)通过这些作用的组合，d)或与HO1相关的其他未知活动。3.确定持续暴露于低水平的NO是否会对毒性NO挑战产生持续的抵抗。研究IAR的窗口是否可以扩大具有明显的治疗意义。 与公共卫生相关：细胞损伤和由自由基引起的死亡，一氧化氮(NO)，见于神经退行性疾病，如多发性硬化症(MS)和肌萎缩侧索硬化症(ALS)，以及急性损伤，如中风、脑创伤和脊髓损伤。低剂量的NO可诱导基因产物--血红素加氧酶1(HO1)的产生，该基因产物可保护神经元免受高剂量NO引起的细胞损伤，这项资助的目的是找出其机制。对HO1介导的通路的解剖和操纵将有助于我们确定治疗或预防NO介导的损伤的治疗靶点，从而减轻MS、ALS和CNS的损伤。

DESCRIPTION (provided by applicant): Nitric oxide (NO) is a free radical that is utilized by the CNS for normal physiological functions. However, when it is released in inappropriate locations or in excess, NO results in the formation of nitrotyrosine, a marker for NO-mediated damage, and massive cell death. Cell death and nitrotyrosine formation are seen in spinal cord injury and in neurodegenerative diseases such as Alzheimer's, multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS). Interestingly, motor neuron cells pretreated with sub-toxic doses of NO gain significant resistance to normally toxic doses of NO, a phenomenon we termed induced adaptive resistance (IAR). Furthermore, when exposed to toxic doses of NO, adapted cells have little to no nitrotyrosine formation. IAR is dependent on the heme metabolizing enzyme, heme oxygenase 1 (HO-1). The overall aim of this grant is elucidate and dissect the IAR phenomenon and by so doing, begin to understand native resistance mechanisms in the CNS that can be utilized to protect neurons against NO-mediated damage seen in CNS injury and neurodegenerative disease. The aims are: 1. To determine whether NO-induced cell death is due to the associated nitration of cellular proteins. If so then a) blocking 3NY formation should also block killing by NO and b) the nitration of cellular proteins, in the absence of NO treatment, should be sufficient to kill cells. 2. To determine how HO1 induces adaptive resistance to NO. Potentially, HO1 may induce adaptive resistance to NO by a.) lowering intracellular concentration of heme, b.) by the generation of heme metabolites such as bilirubin and carbon monoxide, c) by the combination of these actions, d) or by some other unknown activity associated with HO1. 3. To determine whether continuous exposure to low levels of NO will give continuous resistance to toxic NO challenge. An investigation of whether the window of IAR can be expanded has obvious therapeutic implications. PUBLIC HEALTH RELEVANCE: Cell damage and death from the free radical, nitric oxide (NO), is seen in neurodegenerative diseases such as multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS), as well as in acute injuries, such as stroke, brain trauma and spinal cord injury. Low dose NO induces production of a gene product, heme oxygenase 1 (HO1), which protects neurons against NO-mediated cell damage from high doses of NO and the aims of this grant are to find out how. The dissection and manipulation of the HO1- mediated pathway by which low dose NO lends protection against out of context or high dose NO will help us identify therapeutic targets for the treatment or prevention of NO mediated damage and thus mitigate the effects of MS, ALS and CNS injury.