Microglial impact on remyelination

小胶质细胞对髓鞘再生的影响

• 批准号: 10357946
• 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/10357946
• 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.

项目概要 增强髓鞘再生是多发性脱髓鞘患者恢复脑功能的关键策略 硬化症（MS）患者；然而，尽管共同努力，刺激多发性硬化症大脑髓鞘再生的能力 一直难以捉摸。虽然促进少突胶质细胞前体分化的信号通路 已经确定，正在研究的实验环境并没有复制限制机制 MS 特异性炎症性中枢神经系统损伤后的髓鞘再生。当前的提案建立在我们的新模型的基础上 使用多发性硬化症患者产生的致病性重组抗体 (rAb) 进行脱髓鞘/髓鞘再生治疗。 髓磷脂特异性 MS rAb 在体内和离体中直接介导补体介导的脱髓鞘作用，所有这些 与小胶质细胞激活相关的自发修复。然而，脱髓鞘外植体 持续暴露于髓磷脂特异性 MS rAb 无法激活小胶质细胞，并且少突胶质细胞成熟缓慢 抑制。类似地，rAb 介导的脱髓鞘作用后小胶质细胞的靶向清除会被阻断 少突胶质细胞成熟阻止活性髓鞘再生。使用单细胞 RNASeq (scRNASeq) 从髓鞘再生外植体中分离出小胶质细胞，我们鉴定了转录上不同的小胶质细胞亚群，它们是 与成功或失败的髓鞘再生有关。因此，我们假设小胶质细胞信号至关重要 用于从早期髓鞘形成到活跃髓鞘形成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髓鞘再生。测试 根据我们的假设，我们提出了三个互补的具体目标。在目标 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 那些在毒素介导的脱髓鞘中看到的。丘脑内或胼胝体注射髓磷脂特异性 MS rAb 加 HC 将与药理学小胶质细胞耗竭和慢性 施用 MS rAb 来验证小胶质细胞反应对完整神经髓鞘再生的影响 系统。溶血卵磷脂诱导的脱髓鞘作用后将进行类似的研究，脱髓鞘作用非常明显 小胶质细胞激活和髓鞘再生的不同时间过程。在目标 2 中，我们将研究 使用活体成像进行脱髓鞘、小胶质细胞反应和少突胶质细胞原位再生 皮质脱髓鞘。这种对髓磷脂损失、小胶质细胞激活和髓鞘再生的实时分析将是 与铜宗介导的脱髓鞘作用相比。最后，在目标 3 中，我们将调查 小胶质细胞利用髓鞘脱髓鞘的离体小脑切片影响髓鞘再生的机制 特异性 rAb 加人补体 (HC)。我们将重点研究几个小胶质细胞基因的作用 通过 scRNASeq 鉴定预计会促进或损害髓鞘再生。白质外观正常 对具有不同程度脱髓鞘和髓鞘再生的 MS 病变组织进行评估，以确定 功能重要的小胶质细胞亚群的丰度和定位。这些研究的结果将 为控制炎症损伤后髓鞘再生的新机制提供见解。此外， 获得的知识可能会确定新的治疗方法，从而产生具有临床意义的髓磷脂 维修。