Microglial impact on remyelination

小胶质细胞对髓鞘再生的影响

• 批准号: 10175074
• 负责人: Jeffrey L Bennett
• 金额: $ 48.74万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-15 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10175074
• 关键词: Aftercare; Animal Model; Antibodies; Antibody Therapy; Autoantibodies; Brain; CCL2 gene; Cells; Cerebrospinal Fluid; Chronic; Clinical; Complement; Corpus Callosum; Cuprizone; Data; Demyelinations; Exposure to; Gene Expression Profile; Genes; Genetic Transcription; Human; IGF1 gene; Image; Immune response; Immunoglobulin G; Impairment; In Situ; Inflammatory; Infusion procedures; Injections; Injury; Investigation; Knowledge; Lesion; Lysophosphatidylcholines; Mediating; Microglia; Microinjections; Modeling; Monitor; Multiple Sclerosis; Multiple Sclerosis Lesions; Myelin; Natural regeneration; Nervous System Physiology; Nervous System Trauma; Nervous system structure; Neuraxis; Neuroglia; Neurologic; Oligodendroglia; Pathogenicity; Pharmacology; Plasma Cells; Process; Production; Recombinant Antibody; Recovery; Role; Signal Pathway; Signal Transduction; Slice; Testing; Thalamic structure; Time; Tissues; Toxin; Visualization; central nervous system demyelinating disorder; central nervous system injury; clinically relevant; disability; functional restoration; glycoprotein NMB; gray matter; in vivo; in vivo Model; in vivo imaging; injury and repair; insight; intravital imaging; multi-photon; multiple sclerosis patient; novel; novel therapeutic intervention; oligodendrocyte precursor; prevent; remyelination; repaired; response; selective expression; single-cell RNA sequencing; therapy design; white matter

PROJECT SUMMARY Enhancing remyelination is a critical strategy for restoring brain function after demyelination in multiple sclerosis (MS) patients; however, despite concerted efforts, the ability to stimulate remyelination in MS brain has remained elusive. While signaling pathways that promote oligodendrocyte precursor differentiation have been identified, the experimental milieux under investigation do not replicate the mechanisms limiting remyelination following MS-specific inflammatory CNS injuries. The current proposal builds on our new models of demyelination/remyelination using pathogenic recombinant antibodies (rAbs) generated from MS patients. Myelin-specific MS rAbs direct complement-mediated demyelination in vivo and ex vivo, all of which spontaneously repair in association with microglial activation. However, demyelinated explants that are continuously exposed to myelin-specific MS rAb fail to activate microglia, and oligodendrocyte maturation is inhibited. Similarly, targeted depletion of microglia following rAb-mediated demyelination blocks oligodendrocyte maturation preventing active remyelination. Using single cell RNASeq (scRNASeq) on microglia isolated from remyelinating explants, we identified transcriptionally distinct microglial subsets that are associated with successful or failed remyelination. Hence, we hypothesize that microglial signals are critical for oligodendrocyte responses during the transition from early myelinating to actively myelinating oligodendrocyte, and myelin-specific MS autoantibody modulates these signals to arrest remyelination. To test our hypothesis, we propose three complementary specific aims. In Aim 1, we will evaluate microglial and oligodendrocyte responses in in vivo models of MS rAb-mediated demyelination and compare those responses to those seen in toxin-mediated demyelination. Intrathalamic or corpus callosum injection of myelin-specific MS rAb plus HC will be performed in conjunction with pharmacologic microglial depletion and chronic administration of MS rAb to validate the impact of microglial responses on remyelination in the intact nervous system. Comparable studies will be done following lysolecithin-induced demyelination, which has a very different time course of microglial activation and remyelination. In Aim 2, we will study the dynamics of demyelination, microglial responses and oligodendrocyte regeneration in situ using intravital imaging following cortical demyelination. This real-time analysis of myelin loss, microglial activation and remyelination will be compared to that seen following cuprizone-mediated demyelination. Finally, in Aim 3, we will investigate the mechanisms by which microglia impact remyelination using ex vivo cerebellar slices demyelinated with myelin- specific rAb plus human complement (HC). We will focus on investigating the role of several microglial genes identified by scRNASeq that are expected to promote or impair remyelination. Normal appearing white matter and MS lesion tissue with varying degrees of demyelination and remyelination will be evaluated to determine the abundance and localization of functionally-important microglial subsets. The results of these studies will provide insights into novel mechanisms controlling remyelination after inflammatory injury. In addition, the knowledge gained may identify novel therapeutic approaches that will result in clinically-meaningful myelin repair.

项目摘要 增强再髓式是恢复多个脱髓鞘后大脑功能的关键策略 硬化症（MS）患者；但是，尽管努力协同，但刺激MS大脑的再髓的能力 仍然难以捉摸。而促进少突胶质细胞前体分化的信号通路具有 已确定，正在研究的实验环境不会复制限制机制 MS特异性炎症性中枢神经系统损伤后的再髓。当前的建议建立在我们的新模型上 使用MS患者产生的致病性重组抗体（RAB）的脱髓鞘/再髓性。 髓磷脂特异性MS RABS直接补体介导的脱髓鞘在体内和ex Vivo中，所有这些 与小胶质激活相关的自发修复。但是，脱髓鞘的外植体 连续暴露于髓磷脂特异性MS RAB无法激活小胶质细胞，而少突胶质细胞的成熟为 抑制。同样，在Rab介导的脱髓鞘块后，小胶质细胞的靶向耗竭 少突胶质细胞的成熟，可预防主动透明。使用单个单元格RNaseq（scrnaseq） 从再髓外植体中分离出的小胶质细胞，我们确定 与成功或失败的再髓关系有关。因此，我们假设小胶质信号很关键 从早期髓鞘过渡到积极髓鞘的过渡期间的少突胶质细胞反应 少突胶质细胞和髓磷脂特异性MS自身抗体调节这些信号以阻止再髓样。测试 我们的假设，我们提出了三个补充特定目标。在AIM 1中，我们将评估小胶质细胞和 MS Rab介导的脱髓鞘的体内模型中的少突胶质细胞反应并比较这些反应 在毒素介导的脱髓鞘中看到的人。髓鞘或callosum callosum注射髓磷脂特异性MS Rab Plus HC将与药理学小胶质细胞耗竭和慢性一起进行 给予MS RAB以验证小胶质细胞反应对完整神经中的再髓的影响 系统。在溶血石诱导的脱髓鞘之后，将进行可比的研究 小胶质激活和再髓的不同时间过程。在AIM 2中，我们将研究 脱髓鞘，小胶质细胞反应和少突胶质细胞再生，使用静脉内成像以下 皮质脱髓鞘。对髓磷脂损失，小胶质细胞激活和透明的实时分析将是 与丘比酮介导的脱髓鞘之后相比。最后，在AIM 3中，我们将调查 小脑切片用髓磷脂脱髓鞘脱髓鞘的小胶质细胞撞击透明的机制 特定的Rab Plus人类补体（HC）。我们将专注于研究几个小胶质基因的作用 由scrnaseq确定，预计会促进或损害再生。正常出现白质 将评估具有不同程度的脱髓鞘和再生的MS病变组织以确定 功能至关重要的小胶质细胞集的丰度和定位。这些研究的结果将 提供有关控制炎症性损伤后控制透明度的新机制的见解。另外， 获得的知识可能会确定新型的治疗方法，这将导致临床上敏感的髓磷脂 维修。