A human neuron/oligodendrocyte co-culture model to study myelination

研究髓鞘形成的人类神经元/少突胶质细胞共培养模型

• 批准号: 8323029
• 负责人: BRIDGET SHAFIT-ZAGARDO
• 金额: $ 25.01万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8323029
• 关键词: Adhesions; Agonist; Animal Model; Area; Axon; Biological Models; Caliber; Cannabinoids; Cell Maturation; Cells; Chronic Disease; Coculture Techniques; Confocal Microscopy; Data; Demyelinations; Disease; Electron Microscopy; Electrons; Goals; Growth; Growth Factor Receptors; Human; Human Activities; Image; In Vitro; Incubated; Length; Maintenance; Membrane; Methods; Microscopic; Microscopy; Modeling; Multiple Sclerosis; Myelin; Myelin Proteins; Myelin Sheath; Nature; Neurons; Oligodendroglia; Pathology; Patients; Process; Proteins; RXR; Recombinants; Rodent; Role; Rotation; Sagittaria; Series; Signal Transduction; Signaling Molecule; Site; Spinal Ganglia; System; Testing; Therapeutic; Time; alitretinoin; cannabinoid receptor; in vitro Model; insight; myelination; neurofilament; novel; oligodendrocyte precursor; parallel processing; precursor cell; protein expression; receptor; receptor expression; remyelination

DESCRIPTION (provided by applicant): A pivotal issue for patients with multiple sclerosis (MS) is the long-term survival and integrity of neurons and oligodendrocytes, and maintenance of the myelin sheath. Over time, the progressive nature of MS results in a chronic disease course with permanent axonal damage, neuronal and oligodendrocyte loss, demyelination, and diminished re-myelination. Studies in animal models have proved useful in exploring disease mechanisms and in testing therapeutic paradigms, however model systems using cells of human origin are clearly essential for the identification of proteins required to maintain and enhance myelination in humans. In this proposal, our goal is to use our human co-culture system of dorsal root ganglia and oligodendrocyte precursor cells (OPC) to determine the extent to which factors proposed to enhance myelination in rodents function similarly in the human system. We will expand upon our preliminary data that show that growth-arrest specific protein 6 (gas6) enhances OPC maturation and axon ensheathment, and test whether in combination with gas6, the activation of cannabinoid receptors, or the retinoid X receptor (RXR) can promote further the maturation and myelinating activity of human OPC in vitro. In Specific Aim 1, we will continue to examine the ability of gas6 to enhance axonal ensheathment, and examine by immunofluorescent microscopy and electron microscopy the extent to which myelin proteins are expressed in the developing myelin sheath, whether compact myelin is achieved over time, and the correlation between axonal diameter and myelin sheath formation. In Specific Aim 2A, we will examine the contribution of agonists and antagonists of cannabinoid (CB) receptors plus and minus gas6 to determine whether CB2 agonists enhance myelination and aid in myelin compaction. In Specific Aim 2B, we will determine whether 9-cis-retinoic acid signaling through the RXR? receptor enhances myelination. These studies will help to define factors required for human myelination. Analysis of growth factors, receptor agonists and signaling molecules will determine the potential role of these factors in enhancing human myelination with a focus on the factors required for compact myelin formation in a human model system. PUBLIC HEALTH RELEVANCE: We propose to use a novel human neuronal/oligodendrocyte co-culture model to study factors required by human OLs to form myelin. A particular focus of these studies will be to examine the contribution of growth-arrest specific protein 6 (gas6) to oligodendrocyte maturation and myelin protein expression. In addition, we will examine the role of cannabinoid receptors and retinoid X receptors on myelin protein expression and the role of specific agonists and antagonists in enhancing myelination. To our knowledge, our system is the only completely human co-culture model in which mature myelin synthesizing proteins are expressed, and in which OLs are shown to wrap axons by electron microscopy.

描述（由申请人提供）：多发性硬化症（MS）患者的关键问题是神经元和少突胶质细胞的长期存活和完整性，以及髓鞘的维持。随着时间的推移，多发性硬化症的进展性导致慢性病程，伴有永久性轴突损伤、神经元和少突胶质细胞丢失、脱髓鞘和再髓鞘形成减少。动物模型研究已被证明在探索疾病机制和测试治疗范例方面是有用的，然而，使用人类来源细胞的模型系统对于鉴定维持和增强人类髓鞘形成所需的蛋白质显然是必不可少的。在这项提议中，我们的目标是使用我们的人类背根神经节和少突胶质前体细胞（OPC）共培养系统来确定在何种程度上提出的增强啮齿动物髓鞘形成的因素在人类系统中具有类似的功能。我们将扩展我们的初步数据，表明生长抑制特异性蛋白6 （gas6）促进OPC成熟和轴突嵌套，并测试是否与gas6联合，大麻素受体的激活，或视黄醇X受体（RXR）可以进一步促进体外人OPC的成熟和髓鞘活性。在Specific Aim 1中，我们将继续研究gas6增强轴突成鞘的能力，并通过免疫荧光显微镜和电镜检查髓鞘蛋白在发育中的髓鞘中的表达程度，髓鞘是否随着时间的推移而致密化，以及轴突直径与髓鞘形成之间的相关性。在Specific Aim 2A中，我们将检查大麻素（CB）受体+和- gas6的激动剂和拮抗剂的作用，以确定CB2激动剂是否增强髓鞘形成并帮助髓鞘压实。在特异性Aim 2B中，我们将确定9-顺式维甲酸信号是否通过RXR？受体增强髓鞘形成。这些研究将有助于确定人类髓鞘形成所需的因素。对生长因子、受体激动剂和信号分子的分析将确定这些因素在增强人类髓鞘形成中的潜在作用，重点关注人类模型系统中致密髓鞘形成所需的因素。