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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.

灰白质细胞外线粒体转移改善感觉运动和运动功能卒中后认知功能障碍线粒体功能对于维持中枢神经系统(CNS)的细胞内环境平衡至关重要。它现在人们认识到线粒体出人意料地被释放并在细胞之间转移。然而，人们对潜在的机制仍然知之甚少。在这份提案中，我们将调查翻译后 O-GlcNAc作为细胞外线粒体功能的关键元件的修饰和转移介导卒中后血脑屏障恢复、少突胶质细胞保护和神经可塑性。我们的初步数据表明：(I)内质网-高尔基体转运可能调节线粒体蛋白O-GlcN酰化，(Ii) O-GlcN酰化线粒体转移对神经元、少突胶质细胞前体细胞和内皮细胞的保护作用阻断DJ1 O-GlcN酰化可能增加线粒体糖基化和降解，从而降低细胞外线粒体的保护能力，(Iv)水平线粒体O-GlcNAc-DJ1可能与卒中后功能恢复呈正相关，与功能恢复呈负相关与衰老相关，(V)剖析这些机制的方法(分子工具、光学成像、电生理学、SnRNA-seq等)在我们的合作实验室中是可行的。根据我们的试验数据，我们假设损伤相关的分子模式(DAMP)或CD31信号 O-GlcNAc支持上调反应性星形胶质细胞CD38及CD38驱动的翻译后修饰细胞外线粒体功能加速线粒体活性转移介导的神经血管重塑和重新髓鞘形成，从而改善感觉运动障碍和中风后痴呆。我们有三个具体目标。在目标1中，我们将剖析O-GlcNAc修饰的线粒体释放的机制星形胶质细胞与神经元或脑内皮细胞或OPC/少突胶质细胞之间的转移。在目标2中，我们将研究年轻人、老年人和男性灰质和白质中O-GlcNacylated线粒体的转移或局灶性脑缺血后雌性小鼠。在目标3中，我们将使用函数增益和损耗实验来修改线粒体蛋白O-GlcN酰化，并评估体内感觉运动和认知结果。我们的实验将利用分子工具，包括药理激活剂/抑制剂的组合，位点- 定向突变和AAV诱导的线粒体标记评估星形胶质细胞特异性线粒体调职。将进行单核RNA序列以全面定位脑内灰质和白质的转录本。卒中。翻译相关性将通过活体成像、电生理和长期实验进行评估卒中后的结局。这个项目应该定义一种新的线粒体转移机制，在灰色和白质，有望为促进感觉运动恢复和改善后遗症提供新的途径中风血管相关性痴呆。