Using NAD+ precursor for treatment of global cerebral ischemia

总结： 由于心脏骤停或中风引起的缺血性脑损伤是最复杂的病理生理学之一， 流程.为了成功治疗缺血性脑损伤，需要靶向几种细胞通路， 脑内的细胞类型。 NAD+是参与多种生物能量反应及其降解的重要辅因子 局部缺血后导致病理性细胞代谢和能量产生抑制。大多数 的细胞NAD+通过补救途径重新合成，其中烟酰胺单核苷酸（NMN） 转化为NAD+。我们最近的研究表明，NMN的管理显着 改善短暂性全脑缺血后的缺血性脑损伤。此外，NMN 治疗抑制缺血后NAD+ catalysts，减少聚ADP-核糖的产生，并逆转 过度的线粒体碎裂最后，缺血诱导的线粒体变化 蛋白乙酰化在注射NMN的动物中受到抑制。本拟议项目的总体目标是 开发最有效的治疗策略，利用NMN，将大大减少缺血性脑 损伤并表征其神经保护机制。我们假设NMN 由于其多靶点效应，在缺血后给药将显著抑制神经变性 以及改善线粒体功能和细胞生物能量学的能力。 具体目标1集中在NMN诱导的蛋白乙酰化机制，调节 线粒体动力学和功能。Sirt 1和Sirt 3依赖性去乙酰化在 线粒体融合和分裂将使用我们的转基因动物模型来确定， 同时表达线粒体靶向的eYFP和Sirt 1或Sirt 3。SIRT 1和SIRT 3 敲除动物将用作抑制脱乙酰酶活性的工具。具体目标2： 在小鼠全脑缺血后进行NMN给药的时间依赖性研究。我们 将使用无偏的神经元细胞死亡的体视学定量，多个认知测试将 以评估治疗效果。恢复期将从7天到6天不等 缺血性损伤后3个月。在具体目标3中，我们将确定NMN的神经保护作用。 通过使用目标2中确定的最具神经保护性的方案对雌性和老年动物进行治疗。 这项工作的意义在于，它提出将NMN鉴定为保护性化合物， 这将显著影响NAD+前体作为急性胰腺炎的治疗化合物的临床应用。 脑损伤，并可能揭示神经保护的新靶点。

Summary: Ischemic brain damage due to cardiac arrest or stroke is one of the most complex pathophysiologic processes. To successfully treat ischemic brain injury, one need to target several cellular pathways and cell-type within the brain. NAD+ is an essential cofactor involved in multiple bioenergetic reactions and its degradation after ischemia leads to pathologic cellular metabolism and inhibition of energy production. The majority of cellular NAD+ is re-synthetized via the salvage pathway, where nicotinamide mononucleotide (NMN) is converted to NAD+. Our recent studies demonstrated that administration of NMN dramatically ameliorates ischemic brain injury following transient global cerebral ischemia. Furthermore, NMN treatment inhibited post-ischemic NAD+ catabolism, reduced poly-ADP-ribose generation, and reversed the excessive mitochondrial fragmentation. Finally, the ischemia-induced changes in mitochondrial protein acetylation were inhibited in NMN injected animals. Overall goal of this proposed project is to develop the most effective treatment strategy utilizing NMN that will dramatically reduce ischemic brain damage and characterize the mechanism of its neuroprotection. We hypothesize that NMN administration after ischemia will significantly inhibit neurodegeneration due to its multi-targeted effect and ability to improve mitochondrial functions and cellular bioenergetics. Specific Aim 1 is focused on the NMN-induced protein acetylation mechanisms that modulate mitochondrial dynamics and function. The role of Sirt1 and Sirt3 dependent deacetylation in mitochondrial fusion and fission will be determined using our transgenic animal models that concomitantly express mitochondria targeted eYFP and Sirt1 or Sirt3. Additionally, SIRT1, and SIRT3 knockout animals will be used as tools for inhibition of deacetylase activity. In specific Aim 2, we will perform time-dependent studies of NMN administration following global cerebral ischemia in mice. We will use unbiased stereological quantification of neuronal cell death, and multiple cognitive tests will be performed to assess the efficacy of treatment. The recovery periods will vary from 7 days up to 6 months after ischemic insult. In specific Aim 3, we will determine the neuroprotective effect of the NMN treatment by using the most neuroprotective protocol determined in Aim 2 on female and aged animals. The significance of this work is that it proposes to identify NMN as a protective compound that will significantly impact the clinical application of NAD+ precursors as therapeutic compounds for acute brain injury and potentially reveal new targets for neuroprotection.