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Reprogramming proinflammatory microglia by restoring mitochondrial function

通过恢复线粒体功能重新编程促炎性小胶质细胞

基本信息

批准号：
10201784
负责人：
BRIAN M POLSTER
金额：
$ 60.34万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10201784
关键词：
2-oxoglutarate 3-dioxygenase proline Affective Animal Model Antioxidants Attenuated Binding Brain Brain Injuries Cell Fraction Cells Chronic Clinical Cognitive Cognitive deficits Complex Consumption Contusions Data Dementia Drug usage Encephalitis Enzymes Exhibits Female Flow Cytometry Fluorescence Gene Expression Genetic Glycolysis Goals HMGB1 Protein Histopathology Hydroxylation Hypoxia Immune Impairment In Vitro Incubated Individual Inflammatory Injury Interferons Interleukin-1 Interleukin-1 beta Intervention Knock-out Lysosomes Mechanics Metabolic Methods Microglia Mitochondria Modeling Molecular Monitor Motor Mus Nerve Degeneration Neurologic Neurologic Deficit Nitric Oxide Outcome Oxidative Phosphorylation Oxidative Stress Oxygen Oxygen Consumption Pathogenicity Pathway interactions Pharmaceutical Preparations Phenotype Positron-Emission Tomography Process Production Proline Proteins Proteomics Quality Control Reporter Reporting Research Role Severities Signal Transduction Sorting - Cell Movement Source TBI treatment TLR4 gene Testing Traumatic Brain Injury Woman base behavior test brain cell clinically relevant controlled cortical impact cytokine disability effective therapy efficacy evaluation experimental study idebenone in vitro Model in vivo male men mitochondrial dysfunction mouse model neuroprotection novel novel strategies prevent progressive neurodegeneration response response to injury restoration sex translational study

项目摘要

Traumatic brain injury (TBI) is a major source of long-term disability and dementia. Microglia, long-lived immune cells of the brain, activate to multiple reactive states in response to injury. The extent of pro-inflammatory activation correlates with the severity of neurological impairments, suggesting that unresolved activation is pathogenic. The proposed research is highly significant because it will evaluate a clinically safe intervention to ameliorate harmful TBI-induced microglial activation that may contribute to dementia and other chronic neurological deficits following TBI. Our data suggest that a metabolic shift from oxidative phosphorylation to glycolysis during pro-inflammatory activation involves impairment to the lysosomal turnover of damaged mitochondria by mitophagy, which is followed by Complex I subunit degradation. Idebenone restores oxygen consumption by damaged mitochondria and attenuates pro-inflammatory nitric oxide and interleukin-1beta production. The restoration of oxygen consumption by idebenone decreases intracellular oxygen concentration. We found that simply lowering oxygen concentration by incubating cells under hypoxia prevents Complex I degradation and mitophagy impairment. Unexpectedly, antioxidants failed to yield similar rescue, suggesting a role for oxygen that is independent of oxidative stress. Using proteomics, we discovered a marked accumulation of prolyl 3-hydroxylase 2 (P3H2) in a mitochondria/lysosome-enriched cell fraction. P3H2 gene expression within the brain is microglia-specific, and our preliminary data show elevated P3H2 in mouse peri-contusional cortex after TBI. P3H2 enzyme uses oxygen as a substrate for proline hydroxylation of target proteins. Idebenone may suppress the activity of P3H2 by decreasing oxygen availability, preventing P3H2 from post-translationally modifying mitochondria or lysosome proteins involved in quality control. This study will test the central hypothesis that idebenone suppresses TBI-induced microglial activation, chronic neurodegeneration, and cognitive deficits by reversing P3H2-dependent inhibition of mitophagy. The following specific aims employ state-of-the-art mouse models to monitor microglial mitochondrial turnover and include novel methods to sort and study microglia ex vivo based on mitochondrial function. The experiments in Aim 1 will test the prediction that idebenone rescues mitophagy in pro-inflammatory microglial cells by decreasing intracellular oxygen concentration, thereby inhibiting P3H2 activity. Using both male and female mice and considering sex as a variable, the experiments in Aim 2 will test the prediction that idebenone or genetic P3H2 knockout ameliorates TBI-induced pro-inflammatory microglia accumulation and neurological deficits by rescuing microglial mitophagy. Positive outcomes will support translational studies of idebenone to treat TBI-induced dementia, with the potential to help millions of men and women living with the devastating consequences of TBI.

创伤性脑损伤（TBI）是长期残疾和痴呆的主要原因。小胶质细胞，寿命长大脑的免疫细胞会激活多种反应状态以应对损伤。促炎程度激活与神经损伤的严重程度相关，表明未解决的激活是致病的。拟议的研究非常重要，因为它将评估临床安全干预措施改善 TBI 诱导的有害小胶质细胞活化，可能导致痴呆和其他慢性病 TBI 后的神经功能缺损。我们的数据表明，代谢从氧化磷酸化转变为促炎激活过程中的糖酵解涉及受损细胞的溶酶体周转受损线粒体通过线粒体自噬，随后发生复合物 I 亚基降解。艾地苯醌恢复氧气被受损的线粒体消耗并减弱促炎一氧化氮和白介素-1β 生产。艾地苯醌恢复耗氧量，降低细胞内氧浓度。我们发现，简单地通过在缺氧条件下培养细胞来降低氧气浓度可以防止复合物 I 降解和线粒体自噬损伤。出乎意料的是，抗氧化剂未能产生类似的救援作用，这表明氧气的作用与氧化应激无关。使用蛋白质组学，我们发现了明显的积累富含线粒体/溶酶体的细胞组分中脯氨酰 3-羟化酶 2 (P3H2) 的含量。 P3H2基因表达大脑是小胶质细胞特异性的，我们的初步数据显示小鼠挫伤周围皮质中的 P3H2 升高 TBI 后。 P3H2 酶使用氧作为靶蛋白脯氨酸羟基化的底物。艾地苯醌可能通过减少氧气利用率来抑制 P3H2 的活性，防止 P3H2 翻译后修饰参与质量控制的线粒体或溶酶体蛋白。这项研究将测试中央艾地苯醌抑制 TBI 诱导的小胶质细胞激活、慢性神经退行性变的假设通过逆转 P3H2 依赖性线粒体自噬抑制来改善认知缺陷。以下具体目标采用最先进的小鼠模型来监测小胶质细胞线粒体周转，并包括分类和分类的新方法基于线粒体功能离体研究小胶质细胞。目标 1 中的实验将检验以下预测：艾地苯醌通过减少细胞内氧来拯救促炎性小胶质细胞的线粒体自噬浓度，从而抑制 P3H2 活性。使用雄性和雌性小鼠并将性别视为变量，目标 2 中的实验将测试艾地苯醌或基因 P3H2 敲除改善的预测 TBI 通过拯救小胶质细胞线粒体自噬诱导促炎性小胶质细胞积聚和神经功能缺损。积极的结果将支持艾地苯醌治疗 TBI 诱发的痴呆症的转化研究，帮助数百万遭受 TBI 破坏性后果的男性和女性。