Omega 3 fatty acids, acute neuroprotection via mitochondria

项目摘要 新生儿中风和缺氧缺血性（HI）脑损伤仍然是终身神经系统疾病的主要原因。 残疾。目前唯一被接受的方法是缺血后低温。然而，没有确切的治疗方法， 这种方法的机制是已知的。越来越多的实验证据表明，omega-3（n-3） 脂肪酸（FAs）及其生物活性代谢物保护发育中的脑免受HI-再灌注损伤。使用 甘油三酯乳剂，我们发现二十二碳六烯酸（DHA）提供了强大的神经保护，对HI 不同啮齿动物模型中的脑损伤。这种神经保护作用与DHA含量的增加有关， 脑线粒体，在再灌注中保留线粒体功能，并伴有显著的（3至15倍） DHA代谢物、神经保护素D1（NPD 1）和其他专门的促消退介质（SPM）升高， 特别是在缺血的大脑中。我们推断，DHA的有益作用可能是通过其 代谢物。在测试的SPM中，只有NPD 1显著减弱小鼠中的HI脑损伤。管理 NPD 1还抑制了Ca 2+诱导的线粒体膜通透性转换孔的激活 （mPTP）和凋亡死亡途径。在此更新申请中，我们 假设DHA衍生生物活性介质NPD 1 a）修饰基质线粒体膜 减弱mPTP活化的性质，B）阻止线粒体外膜与BAX的相互作用。 这些作用减弱缺血后二次能量衰竭和线粒体凋亡细胞死亡途径， 有助于/解释DHA提供的神经保护机制。这一假设将在三个 明确的目标。目的1：确定外源性NPD 1和DHA在减弱BAX驱动的细胞凋亡中的作用。 HI脑损伤后线粒体细胞死亡。在这个目标中，我们将比较DHA或NPD 1在 限制BAX介导的细胞死亡。考虑到两种化合物都表现出有益的变化， 线粒体功能（Ca 2+缓冲和呼吸（DHA）和Ca 2+缓冲（NPD 1））在HI损伤后，我们 集中于确定外源性DHA及其代谢产物NPD 1的命运，通过追踪氘代2 H10-DHA， 缺血脑和从缺血半球分离的线粒体/线粒体体。目标2：确定 急性注射DHA后脑NPD 1增加的起源及NPD 1和外源性DHA的分布 受伤后）。最后，在目标3中，我们将确定NPD 1或/和DHA在保存 线粒体内膜的完整性，直接测试由线粒体的存在或 没有渗透性转变。我们的建议是一个翻译研究，重点是特定的机制， 利用创新方法对缺血后线粒体进行神经保护， 线粒体通透性转换在二次能量衰竭和细胞损伤中时间作用平移 这些影响是由DHA代谢物的神经保护强度和临床效用定义的，而不是DHA本身。

Project Summary Neonatal stroke and hypoxia-ischemia (HI) brain injury remains a leading cause of a lifelong neurological handicap. The only currently accepted approach is post-ischemic hypothermia. However, no exact therapeutic mechanisms of this approach are known. A growing body of experimental evidence showed that omega-3 (n-3) fatty acids (FAs) and their bioactive metabolites protect developing brain against HI-reperfusion injury. Using triglyceride emulsions we showed that docosahexaenoic acid (DHA) provided strong neuroprotection against HI brain injury in different rodent models. This neuroprotection was associated with increased DHA content in cerebral mitochondria, preserved mitochondrial function in reperfusion and coupled with a dramatic (3 to 15 fold) elevation of DHA metabolites, neuroprotectin D1 (NPD1) and other specialized pro-resolving mediators (SPMs), specifically in the ischemic brain. We have reasoned that beneficial action of DHA may be exerted by its metabolites. Among tested SPMs, only NPD1 significantly attenuated HI brain damage in mice. Administration of NPD1 also suppressed activation of Ca2+ induced mitochondrial membrane permeability transition pore (mPTP) and apoptotic death pathway by blocking BAX-mitochondria interaction. In this renewal application, we hypothesize that DHA-derived bioactive mediator, NPD1 a) modifies the matrix mitochondrial membrane properties that attenuate activation of mPTP, b) prevents mitochondrial outer membrane interaction with BAX. These effects attenuate post-ischemic secondary energy failure and mitochondrial apoptotic cell death pathway, contributing/explaining the neuroprotective mechanisms afforded by DHA. This hypothesis will be tested in three specific aims. Aim 1: To determine the role of exogenous NPD1 and DHA in attenuation of BAX-driven mitochondrial cell death following HI brain injury. In this aim, we will compare the potency of DHA or NPD1 in limiting BAX-mediated cellular death. Given that both compounds demonstrated beneficial changes in mitochondrial functions (Ca2+ buffering and respiration (DHA) and Ca2+ buffering (NPD1)) after HI insult, we were focused on determining the fate of exogenous DHA and its metabolite, NPD1 by tracing deuterated 2H10-DHA in the ischemic brain, and mitochondria/mitoplasts isolated from the ischemic hemisphere. Aim 2: To determine the origin of increased brain NPD1 after acute injection of DHA and distribution of NPD1 and exogenous DHA following HI injury). Finally, in Aim 3, we will determine potential actions of NPD1 or/and DHA on preserving an integrity of the inner mitochondrial membrane, directly testing electrical conductance driven by the presence or absence of permeability transition. Our proposal is a translational study with a focus on specific mechanisms of neuroprotection targeting post-ischemic mitochondria using innovative approaches toward an understanding a temporal role of mitochondrial permeability transition in secondary energy failure and cell injury. Translational impacts are defined by the neuroprotective strength and clinical utility of DHA metabolites, rather than DHA itself.