Extracellular Mitochondria Transfer in Gray and White Matter for Ameliorating Sensorimotor and Cognitive Deficits After Stroke

灰质和白质中的细胞外线粒体转移可改善中风后的感觉运动和认知缺陷

基本信息

批准号：
10571863
负责人：
Kazuhide Hayakawa
金额：
$ 50.3万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-15 至 2027-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10571863
关键词：
Acceleration Aging Allografting Animals Astrocytes Blood - brain barrier anatomy Blood Vessels Brain CRISPR/Cas technology Cell Survival Cells Central Nervous System Cerebral Ischemia Cognitive Cognitive deficits Collaborations Confocal Microscopy Cyclic ADP-Ribose Data Dementia Electrophysiology (science)Elements Endothelial Cells Endothelium Energy Supply Etiology Female Glucose Golgi Apparatus Homeostasis Human In Vitro Infarction Integrins Ischemia Knowledge Label Mammalian Cell Maps Mass Spectrum Analysis Mediating Metabolism Methods Mitochondria Mitochondrial Proteins Modification Molecular Mus Neurologic Neuronal Plasticity Neurons Oligodendroglia Oligonucleotides Outcome Oxygen PARK7 gene PECAM1 gene Pathway interactions Pattern Plasmids Play Post-Translational Protein Processing Process Property Proteins Publishing Rattus Recombinants Recovery Recovery of Function Role Sensorimotor functions Signal Induction Signal Pathway Signal Transduction Site-Directed Mutagenesis Small Interfering RNA Specific qualifier value Stroke Testing Therapeutic Tissues Western Blotting angiogenesis behavioral outcome brain endothelial cell cognitive function cognitive recovery cognitive testing deprivation experimental study extracellular functional improvement gain of function glycation gray matter improved in vivo in vivo imaging inhibitor loss of function male neural growth neurogenesis neurological recovery neuroprotection neurovascular neutralizing antibody novel novel strategies novel therapeutic intervention oligodendrocyte precursor optical imaging pharmacologic post stroke post stroke dementia potential biomarker precursor cell prevent remyelination repaired response single nucleus RNA-sequencing spatiotemporal tool trafficking transcriptome white matter

项目摘要

Extracellular mitochondria transfer in gray and white matter for ameliorating sensorimotor and cognitive deficits after stroke Mitochondrial function is essential for maintaining cellular homeostasis in the central nervous system (CNS). It is now recognized that mitochondria are surprisingly released and transferred between cells. However, the underlying mechanisms remain poorly understood. In this proposal, we will investigate post-translational modification by O-GlcNAc as a critical element for extracellular mitochondrial functionality and the transfer- mediated blood-brain barrier recovery, oligodendrocyte protection and neural plasticity after stroke. Our pilot data suggest that (i) ER-Golgi trafficking may regulate mitochondrial protein O-GlcNAcylation, (ii) transfer of O-GlcNAcylated mitochondria may protect neurons, oligodendrocyte precursors and endothelial cells against oxygen-glucose deprivation, (iii) blockade of DJ1 O-GlcNAcylation may increase mitochondrial glycation and degradation, thus reducing the protective capacity of extracellular mitochondria, (iv) levels of mitochondrial O-GlcNAc-DJ1 may be positively correlated with functional recovery after stroke and negatively correlated with aging, (v) methods to dissect these mechanisms (molecular tools, optical imaging, electrophysiology, snRNA-seq etc) are feasible in our collaborating labs. Based on our pilot data, we hypothesize that damage associated molecular pattern (DAMP) or CD31 signals upregulate CD38 in reactive astrocytes, and CD38-driven post-translational modification by O-GlcNAc supports extracellular mitochondrial functionality that accelerates viable mitochondrial transfer-mediated neurovascular remodeling and remyelination, thus ameliorating sensorimotor deficits and post-stroke dementia after stroke. We have 3 specific aims. In Aim 1, we will dissect mechanisms of O-GlcNAc-modified mitochondrial release and transfer between astrocytes and neurons or brain endothelial cells or OPCs/oligodendrocytes. In Aim 2, we will investigate O-GlcNAcylated mitochondrial transfer in gray matter and white matter in young, older, male or female mice after focal ischemia. In Aim 3, we will use gain- and loss-of-function experiments to modify mitochondrial protein O-GlcNAcylation, and assess sensorimotor and cognitive outcomes in vivo. Our experiments will utilize molecular tools including a combination of pharmacologic activators/inhibitors, site- directed mutagenesis, and AAV-induced mitochondrial labeling to assess astrocyte-specific mitochondria transfer. Single-nuclei RNA seq will be performed to fully map transcriptomes in gray and white matter post- stroke. Translational relevance will be assessed with in vivo imaging, electrophysiology and long-term outcomes post-stroke. This project should define a novel mechanism of mitochondrial transfer in both gray and white matter, and hopefully provide new approaches to accelerate sensorimotor recovery and ameliorate post- stroke vascular-related dementia.

细胞外线粒体在灰色和白质中转移，以改善感觉运动和中风后认知缺陷线粒体功能对于维持中枢神经系统（CNS）中的细胞稳态至关重要。它现在认识到线粒体在细胞之间出奇地释放并转移。但是，潜在的机制仍然很少理解。在此提案中，我们将调查翻译后 O-GLCNAC修饰是细胞外线粒体功能和转移 - 的关键元素中风后介导的血脑屏障恢复，少突胶质细胞保护和神经可塑性。我们的试点数据表明，（i）er-golgi运输可能调节线粒体蛋白O-glcnacylation，（ii） O-Glcnacylated线粒体的转移可能保护神经元，少突胶质前体和内皮抗氧葡萄糖剥夺的细胞，（iii）DJ1 O-Glcnacylation的阻断可能会增加线粒体糖基化和降解，从而降低了细胞外线粒体的保护能力，（iv）水平线粒体O-GLCNAC-DJ1可能与中风后的功能恢复正相关，并负面相关与衰老相关，（v）剖析这些机制的方法（分子工具，光学成像，在我们的协作实验室中，电生理学，SNRNA-SEQ等是可行的。根据我们的试点数据，我们假设损害相关的分子模式（潮湿）或CD31信号在反应性星形胶质细胞中上调CD38，以及通过O-GLCNAC支持的CD38驱动的翻译后修饰细胞外线粒体功能，可加速可行的线粒体转移介导的神经血管重塑和再髓式，从而改善了中风后的感觉运动缺陷和中风后痴呆症。我们有3个具体目标。在AIM 1中，我们将剖析O-GlCNAC修饰的线粒体释放的机制以及星形胶质细胞和神经元或脑内皮细胞或OPC/少突胶质细胞之间的转移。在AIM 2中，我们将研究灰质的O-GlcnacyLated线粒体转移，年轻，年龄较大，男性局灶性缺血后的雌性小鼠。在AIM 3中，我们将使用功能丧失实验来修改线粒体蛋白O-Glcnacylation，并在体内评估感觉运动和认知结果。我们的实验将利用分子工具，包括药理学激活剂/抑制剂，部位 - 定向诱变和AAV诱导的线粒体标记，以评估星形胶质细胞特异性线粒体转移。将进行单核RNA SEQ，以完全绘制灰色和白质的转录组 - 中风。翻译相关性将通过体内成像，电生理和长期评估结果后的结果。该项目应定义灰色和灰色和线粒体转移的新机制白质，希望提供新的方法来加速感觉运动恢复并改善后中风血管相关痴呆。