Mitochondrial Structural and Functional Remodeling in Microglial Activation

 描述(申请人提供)：线粒体分裂促进蛋白Drp1与急性和慢性神经退行性疾病的神经病理学有关。MDVI-1是一种血脑屏障渗透性药物，被认为可以抑制Drp1，在中风、心脏骤停和帕金森病的临床前模型中保护大脑。虽然针对神经元中线粒体分裂-融合平衡的测试正在进行中，但在非神经元脑细胞中针对线粒体分裂的影响几乎完全是针对神经疾病。 未被开发的。我们看到小胶质细胞进入促炎激活状态时，线粒体结构重塑与DRp1依赖的分裂一致，称为M1。从氧化磷酸化到糖酵解的生物能量转换促进了M1的激活，而激活的标志是一氧化氮(NO)和促炎细胞因子的产生。类似于人类脑损伤中的神经炎症状态，M1的激活可以持续几个月到几年，并导致额外的病理。更好地了解M1激活的机制将有助于发现阻止M1小胶质细胞激活的有害后果的新疗法。我们发现，DRP1抑制剂MDIVI-1抑制小胶质细胞NO和肿瘤坏死因子-α的产生，以响应内毒素+干扰素-γ。我们还发现，MDVI-1可以抑制线粒体复合体I产生的活性氧(ROS)，这种抑制作用不依赖于DRP1。重要的是要确定已知的mdivi-1靶点之一或两个，即Drp1和ROS，是否有助于体内M1小胶质细胞的激活，从而代表真正的治疗靶点。这项研究将检验中心假说 在体内，小胶质细胞的激活需要的是Drp1的活性，而不是线粒体的ROS，而 随之而来的细胞生物能量学的转变。脑室注射脂多糖可诱导小鼠小胶质细胞活化。为了研究Drp1的作用，将建立一种新的小胶质细胞特异性可诱导基因敲除小鼠模型。此外，现有的MCAT小鼠异位表达基质靶向、过氧化氢解毒过氧化氢酶将被用来评估线粒体ROS独立于其他ROS的特定需求。在目标1中，我们将确定Drp1是否是内毒素诱导的生物能移和小胶质细胞M1激活所必需的。在目标2中，我们将确定线粒体ROS是否是体内注射内毒素诱导的生物能移和小胶质细胞M1激活所必需的。我们的研究将向确定小胶质细胞线粒体是否可以作为治疗的靶点迈出关键的一步。由于正常的依赖于Drp1的分裂是线粒体运输和神经元质量控制所必需的，因此，最终针对小胶质细胞中的Drp1的策略可能比整体抑制线粒体分裂更可取。我们的长期目标是针对缺血和创伤性脑损伤模型中小胶质细胞激活的线粒体机制，最终希望开发出一种对人类的保护性治疗方法。

 DESCRIPTION (provided by applicant): The mitochondrial fission-promoting protein Drp1 is linked to neuropathology in acute and chronic neurodegenerative disorders. Mdivi-1, a blood-brain barrier permeable drug thought to inhibit Drp1, protects the brain in preclinical models of stroke, cardiac arrest, and Parkinson's disease. While major efforts are underway to test whether targeting the mitochondrial fission-fusion balance in neurons can modify neurodisease, the implications of targeting mitochondrial fission in non-neuronal brain cells are almost entirely unexplored. We see a mitochondrial structural remodeling consistent with Drp1-dependent fission in microglia when they enter a proinflammatory activation state termed M1. A bioenergetic shift from oxidative phosphorylation to glycolysis promotes M1 activation, and activation is marked by production of nitric oxide (NO) and proinflammatory cytokines. Resembling the neuroinflammatory state seen in human brain injury, M1 activation can persist for months to years and result in additional pathology. A better understanding of the mechanisms contributing to M1 activation will help discover novel therapeutics that block the deleterious consequences of M1 microglial activation. We find that the Drp1 inhibitor mdivi-1 suppresses microglial NO and TNF-α production in response to lipopolysaccharide (LPS) ± interferon-γ. We also find that mdivi-1 can inhibit reactive oxygen species (ROS) production by mitochondrial complex I in a Drp1-independent fashion. It is important to determine whether one or both of the known mdivi-1 targets, Drp1 and ROS, contribute to M1 microglial activation in vivo, and thus represent bona fide targets for therapy. This study will test the central hypothesis that in vivo, Drp1 activity but not mitochondrial ROS is required for microglial activation and the accompanying shift in cellular bioenergetics. Microglial activation will be induced in mice by intracerebroventricular administration of LPS. To investigate the role of Drp1, a novel microglia-specific inducible knockout mouse model will be developed. In addition, the existing MCAT mouse ectopically expressing matrix-targeted, H2O2-detoxifying catalase will be used to evaluate a specific requirement for mitochondrial ROS independent of other ROS. In Aim 1, we will determine whether Drp1 is necessary for the bioenergetic shift and microglial M1 activation induced by LPS in vivo. In Aim 2, we will determine whether mitochondrial ROS are required for the bioenergetic shift and microglial M1 activation induced by in vivo LPS injection. Our studies will take a crucial step toward determining whether microglial mitochondria can be targeted for therapy. Since normal Drp1-dependent fission is required for mitochondrial trafficking and quality control in neurons, a strategy specifically targeting Drp1 in microglia may ultimately be more desirable than global inhibition of mitochondrial fission. Our long-term goal is to target mitochondrial mechanisms of microglial activation in models of ischemic and traumatic brain injury, with the ultimate hope of developing a protective therapy for humans.