Toll-Like Receptor-Mediated Photoreceptor Mitochondrial DNA Damage

Toll 样受体介导的光感受器线粒体 DNA 损伤

• 批准号: 7633114
• 负责人: NARSING A RAO
• 金额: $ 40.75万
• 依托单位: DOHENY EYE INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7633114
• 关键词: Adaptor Signaling Protein; Address; Age related macular degeneration; Animals; Apoptosis; Apoptotic; Autoimmune Diseases; Blindness; Bone Marrow; Caspase; Cells; Chimera organism; DNA Damage; Demyelinations; Development; Distant; Event; Generations; Genes; Genus Mycobacterium; Heating; Immune; Immune response; Immunity; Infiltration; Inflammation; Inflammatory; Inner Nuclear Layer; Knock-out; Mediating; Microglia; Mitochondria; Mitochondrial DNA; Modeling; Mus; Myelogenous; Natural Immunity; Neuraxis; Neurons; Organ; Outcome; Oxidative Stress; Oxidative Stress Induction; Pathogenesis; Pathologic Processes; Pathology; Pathway interactions; Photoreceptors; Play; Predisposition; Receptor Activation; Reperfusion Injury; Research Methodology; Retina; Retinal; Retinal Photoreceptors; Role; Site; Stress; Systemic disease; T-Lymphocyte; TLR4 gene; Testing; Therapeutic Intervention; Tissues; Toll-like receptors; Tumor Necrosis Factor Receptor; Up-Regulation; Uveitis; base; chemokine; cytokine; design; ganglion cell; in vivo; killings; knockout animal; neuron apoptosis; novel; public health relevance; receptor; receptor expression; receptor upregulation; retinal damage; transcription factor

DESCRIPTION (provided by applicant): Tissue damage in autoimmune disease has previously been attributed to T-cell and inflammatory cell infiltration and to the cytokines released thereafter. Activation of Toll-like receptors (TLRs) in innate immunity generates similar proinflammatory cytokines as those of adaptive immunity and inflict mitochondrial DNA damage. Such TLRs pathology has been overlooked until recently. Among various systemic diseases, TLRs activation has been implicated in the pathogenesis of CNS demyelination, ischemia/reperfusion injury of the organs, and age-related macular degeneration. In this application, TLR4 induced by heat-killed mycobacteria injected at distant sites initiates neuronal cell oxidative stress and mitochondrial DNA damage in the retina. The TLR4 upregulation was also found to associate primarily with the retinal microglia. Based on these preliminary findings, the following specific aims are proposed: 1. Determine the expression and localization of TLR and TNF-1 and its receptors in the mouse retina with an innate immune response. 2. Determine the in-vivo effects of TLR upregulation and interaction with MyD88 on mitochondrial oxidative stress, using various TLRs, MyD88, and other adaptor proteins knockout animals. 3. Determine the role of bone marrow derived retinal microglia TLR upregulation in the retinal neuronal cell mitochondrial oxidative stress, using bone marrow chimeras that express MyD88 limited to the microglia. 4. Localize oxidative stress-mediated mitochondrial DNA damage and caspases in the retina with upregulation of TLRs and demonstrate augmentation of retinal damage with T-cell-mediated adaptive immune response. The research methods designed are closely adhered to these specific aims. In innate immunity, TLRs, transcription factors (such as NFkB), cytokines (such as TNF-1), and iNOS are upregulated (specific aim 1) and are likely to co-localize to retinal microglia (specific aim 1). However, co-localization with other retinal immune cells will be also tested (specific aim 1). The association of TLRs to microglia will be further substantiated by bone marrow chimeric mice in which the retinal microglia will be bone marrow derived (Specific aim 3). The mitochondrial DNA damage mediated by TLRs activation will be tested by using various wild types and knockouts, including adaptor protein knockouts (specific aim 2). Further, the site of DNA damage and caspases will be sought in various retinal cells (specific aim 4). Once clarified, these results on initial events of TLR activation might have a global implications in autoimmune diseases and specific DNA damage in neuronal cells from TLR4 activation. PUBLIC HEALTH RELEVANCE: Intraocular inflammation, or uveitis, is a leading cause of blindness from retinal photoreceptor cell degeneration. This application deals with the previously overlooked aspect of early photoreceptor damage initiated by the innate immunity alone. By clarifying the mechanism of retinal damage rendered by Toll-like receptor activation, we can advance the design of therapeutic intervention to achieve the desired outcome.

描述（由申请人提供）：自身免疫性疾病中的组织损伤先前已归因于T细胞和炎症细胞浸润以及此后释放的细胞因子。先天免疫中的Toll样受体（TLR）的激活会产生与适应性免疫的促炎细胞因子相似的，并造成线粒体DNA损伤。直到最近，这种TLRS病理学一直被忽视。在各种全身性疾病中，TLRS激活与中枢神经系统脱髓鞘，器官的缺血/再灌注损伤以及与年龄相关的黄斑变性有关。在此应用中，在远处注射的热杀死的分枝杆菌诱导的TLR4引发了视网膜中神经元细胞氧化应激和线粒体DNA损伤。还发现TLR4上调主要与视网膜小胶质细胞相关。基于这些初步发现，提出了以下特定目标：1。确定TLR和TNF-1及其受体在小鼠视网膜中的表达和定位，并具有先天的免疫反应。 2。使用各种TLR，MyD88和其他衔接蛋白基因敲除动物，确定TLR上调和与MyD88对线粒体氧化应激的相互作用。 3。确定骨髓衍生的视网膜小胶质细胞TLR上调在视网膜神经元细胞线粒体氧化应激中，使用表达MyD88的骨髓嵌合体限制于小胶质细胞。 4。在视网膜中定位氧化应激介导的线粒体DNA损伤和caspase，并在TLR上上调，并显示出具有T细胞介导的适应性免疫反应的视网膜损伤增强。设计的研究方法紧密遵守这些特定目的。在先天免疫力中，TLR，转录因子（例如NFKB），细胞因子（例如TNF-1）和INOS被上调（特定目标1），并且可能会共定位于视网膜小胶质细胞（特定目标1）。但是，还将测试与其他视网膜免疫细胞共定位（特定目标1）。 TLR与小胶质细胞的关联将被骨髓嵌合小鼠进一步证实，其中视网膜小胶质细胞将是骨髓衍生的（特定的AIM 3）。 TLRS激活介导的线粒体DNA损伤将通过使用各种野生类型和敲除（包括适配器蛋白敲除）测试（特定AIM 2）。此外，将在各种视网膜细胞中寻求DNA损伤和胱天蛋白酶的部位（特定目标4）。一旦澄清，这些结果是TLR激活的初始事件可能对自身免疫性疾病和TLR4激活中神经元细胞中特定的DNA损伤具有全球影响。公共卫生相关性：眼内炎症或葡萄膜炎是视网膜感光细胞变性引起失明的主要原因。该应用程序涉及仅先天免疫引发的早期光感受器损害的先前被忽视的方面。通过阐明通过Toll样受体激活造成的视网膜损伤的机制，我们可以提高治疗干预的设计，以达到预期的结果。