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Neuroinflammation in Wallerian Degeneration and Regeneration: Neutrophils Play a Primary Role as Phagocytes

华勒变性和再生中的神经炎症：中性粒细胞作为吞噬细胞发挥主要作用

基本信息

批准号：
10447730
负责人：
RICHARD E ZIGMOND
金额：
$ 50.03万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
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：
2020-08-01 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10447730
关键词：
Acute Animal Model Anti-Inflammatory Agents Antibodies Attention Axon Axotomy B-Lymphocytes Blood Brain CCL2 gene CXCL1 gene CXCL2 gene Cells Demyelinating Diseases Demyelinations Disease Disease Progression Distal Electrons Electrophysiology (science)Enzyme-Linked Immunosorbent Assay Excision Experimental Autoimmune Neuritis Ganglia Genes Growth Guillain Barré Syndrome IL8RB gene Immune In Situ Hybridization Inflammatory Injury Interleukin-8B Receptor Knock-out Knockout Mice Label Lesion Measures Modeling Multiple Sclerosis Mus Myelin Myelin Proteins Natural regeneration Nerve Nerve Crush Nerve Degeneration Nerve Regeneration Nervous system structure Neural Conduction Neuraxis Neuroglia Neuroimmune Neurons Neuropathy Neutrophil Infiltration Neutrophilic Infiltrate Oils Pathologic Pathology Peripheral Peripheral Nervous System Peripheral Nervous System Diseases Phagocytes Phagocytosis Pharmacology Phenotype Play Population Process Proteins Public Health Recovery of Function Reporting Role Schwann Cells Spinal Cord Stains T-Lymphocyte Techniques Time Tissues Tolonium chloride Trauma Wallerian Degeneration Western Blotting Wild Type Mouse antagonist base cell type chemokine chemokine receptor experimental study improved indexing injured macrophage monocyte monocyte chemoattractant protein 1 receptor mouse model nerve transection nervous system disorder neuroinflammation neutralizing antibody neutrophil novel receptor regenerative relating to nervous system response sciatic nerve

项目摘要

Understanding the process of neural degeneration is important in order to understand the responses of the nervous system to injury and disease. A common model used in such studies is the distal segment of the sciatic nerve after nerve transection or crush. Following such a lesion, Wallerian degeneration occurs in which the distal segment of the nerve fragments and degenerates, and the resulting axonal and myelin debris are cleared away. Interestingly, while this process occurs rapidly in the peripheral nervous system (PNS), it is extremely slow in the central nervous system (CNS), and, partly because of this, regeneration is generally ineffective in the brain and spinal cord. It is widely believed that inflammatory macrophages (mφs) derived from blood-borne monocytes are required for the phagocytosis of axonal and myelin debris. Immune cells enter injured tissue in response to chemotactic cytokines or chemokines. A major population of monocytes infiltrates into the distal nerve after axotomy in response to the chemokine CCL2, which acts on monocytes via the chemokine receptor CCR2. In mice in which the gene for CCR2 is knocked out, CCR2+ monocytes do not enter the nerve. It was, therefore, very surprising when we found that the clearance of both myelin and axonal protein is normal in Ccr2 knockout mice. We subsequently found that an important reason for the normal clearance appears to be phagocytosis by neutrophils, an immune cell not previously implicated in Wallerian degeneration. In fact, although neutrophil actions in the CNS are beginning to be examined, for example in models of multiple sclerosis, there are almost no studies on their actions in the PNS. Our preliminary evidence using the neutrophil-depleting antibody anti- Ly6G suggests, but does not prove, that neutrophils directly phagocytose and metabolize myelin. We will examine this hypothesis by examining the clearance of myelin proteins by western blotting before and after neutrophil depletion and by co-labeling tissue with Oil Red O, a stain for myelin metabolites, and with cell type specific neutrophils antibodies. We have shown that after axotomy two neutrophil attracting chemokines are induced in the sciatic nerve, CXCL1 and CXCL2. To determine whether these chemokines and their receptor, CXCR2, are involved in neutrophil infiltration into the nerve, we will use neutralizing antibodies, pharmacological antagonists, and knockout mice. We will investigate also whether the involvement of neutrophils in Wallerian degeneration is important for subsequent regeneration. “Wallerian-like” degeneration occurs in several demyelinating neuropathies, and we will examine whether neutrophils play a role in this phenomenon. We will use a mouse model for Guillain Barré syndrome based on our recent finding that neutrophils enter into the sciatic nerve in this model. Following up on our unexpected findings of normal clearance of myelin in Ccr2 knockout mice, our experiments will examine the role of neutrophils in Wallerian degeneration and in a demyelinating disease. Given the known importance of Wallerian degeneration for nerve regeneration due to the removal of myelin proteins, our studies will suggest ways of improving regeneration in the PNS and perhaps in the CNS.

了解神经退化的过程对于了解神经元的反应是很重要的。神经系统损伤和疾病。这种研究中常用的模型是坐骨神经的远段神经切断或挤压后的神经。在这样的病变之后，发生沃勒变性，其中远端的神经节段断裂并退化，并且所产生的轴突和髓鞘碎片被清除。有趣的是，虽然这一过程在外周神经系统（PNS）中发生得很快，但在中枢神经系统中却非常缓慢。中枢神经系统（CNS），部分原因是，再生通常在大脑中无效和脊髓。炎症巨噬细胞（Mφs）来源于血单核细胞，是吞噬轴突和髓鞘碎片所必需的。免疫细胞进入受伤的组织，趋化性细胞因子或趋化因子。单核细胞的主要群体浸润到远端神经后，轴突切断反应于趋化因子CCL 2，其通过趋化因子受体CCR 2作用于单核细胞。在在CCR 2基因被敲除的小鼠中，CCR 2+单核细胞不会进入神经。因此，当我们发现在Ccr 2敲除中髓鞘和轴突蛋白的清除是正常的时，小鼠我们随后发现，正常清除的一个重要原因似乎是由嗜中性粒细胞，一种以前与沃勒变性无关的免疫细胞。事实上，尽管中性粒细胞在中枢神经系统中的作用开始被检查，例如在多发性硬化症的模型中，几乎有没有研究他们在PNS的行为。我们的初步证据是使用嗜中性粒细胞消耗抗体抗- Ly 6 G表明，但不能证明，中性粒细胞直接吞噬和代谢髓鞘。我们将通过用蛋白质印迹法检查在注射前和注射后髓磷脂蛋白的清除率来检验这一假设。中性粒细胞耗竭，并用油红O（一种髓鞘代谢物染色剂）和细胞类型共标记组织特异性中性粒细胞抗体。我们已经表明，轴突切断后，两种中性粒细胞吸引趋化因子，在坐骨神经中诱导，CXCL 1和CXCL 2。为了确定这些趋化因子及其受体， CXCR 2，参与中性粒细胞浸润到神经，我们将使用中和抗体，药理学拮抗剂和基因敲除小鼠。我们还将研究嗜中性粒细胞是否参与沃勒氏病退化对于随后的再生是重要的。“沃勒样”变性发生在几个脱髓鞘性神经病，我们将检查中性粒细胞是否在这种现象中发挥作用。我们将基于我们最近发现中性粒细胞进入坐骨神经，神经在这个模型中。在Ccr 2基因敲除中，我们意外发现髓鞘正常清除小鼠，我们的实验将检查中性粒细胞在沃勒变性和脱髓鞘中的作用。疾病考虑到已知的沃勒变性对于由于去除神经纤维而引起的神经再生的重要性，髓鞘蛋白质，我们的研究将提出改善PNS和可能的CNS再生的方法。