Neuronal regulation of myelination

描述（由申请人提供）：在包括人类在内的脊椎动物中，外周神经系统（PNS）和中枢神经系统（CNS）的快速神经元通信依赖于适当的髓鞘形成。髓鞘形成细胞在PNS是雪旺细胞（SC）和在中枢神经系统是少突胶质细胞（OL）。这些特化的细胞包住神经元过程，从而促进电脉冲的快速传播。PNS髓磷脂在几种CMT疾病中存在缺陷，CMT是最常见的遗传性神经系统疾病之一。中枢神经系统髓磷脂的异常发育导致被称为脑白质营养不良的疾病。我们之前描述了严重的周围神经病变CMT4J，由人类FIG4/SAC3基因突变引起，该基因编码一种进化上保守的脂质磷酸酶，该酶调节沿内溶酶体途径的细胞内囊泡运输。我们研究的主要目的是了解FIG4缺乏破坏髓磷脂形成的分子机制，并在临床前模型中制定CMT4J的治疗策略。Fig4整体表达缺失的突变小鼠（Fig4-/-）表现出CNS和PNS髓磷脂的显著减少、严重震颤和幼年致死。电生理记录显示坐骨神经和视神经的电脉冲传导减慢。令人惊讶的是，通过野生型图4的神经元特异性表达，可以“拯救”Fig4-/-小鼠的髓磷脂缺陷。基于这些观察，我们假设图4的缺失破坏了髓鞘形成所需的神经元特异性信号机制。在特异性目标1和目标2中，我们结合小鼠遗传学和蛋白质组学来鉴定Fig4突变小鼠中被破坏的神经元髓鞘形成信号，并确定体内对Fig4的时间需求。这些实验将为指导髓鞘形成的神经元信号提供新的机制见解。为了模拟人类CMT4J，我们开发了在Fig4-/-背景下普遍表达低水平人类疾病等位基因Fig4- i41t的转基因小鼠（CMT4J小鼠）。这些小鼠表现出与图4-/-小鼠相当的髓鞘退化，但存活到成年后仍具有许多人类疾病的神经系统特征。由于我们已经证明Fig4在神经元中的转基因表达足以驱动髓鞘形成，我们在Specific Aim 3中提出了基因治疗研究。用病毒载体转染CMT4J小鼠的背根神经节神经元（PNS）和视网膜神经节细胞（CNS）表达野生型（图4）。将监测坐骨神经或视神经的髓鞘形成、淋巴结结构和神经传导速度作为疗效指标。通过Fig4基因治疗恢复小鼠髓鞘形成将为髓鞘形成障碍患者提供新的治疗途径。

DESCRIPTION (provided by applicant): In vertebrates, including humans, rapid neuronal communication in the peripheral (PNS) and central nervous system (CNS) is dependent on proper myelination. The myelin-forming cell in the PNS is the Schwann cell (SC) and in the CNS the oligodendrocyte (OL). These specialized cells ensheath neuronal processes and thereby facilitate rapid propagation of electrical impulses. PNS myelin is defective in several types of Charcot-Marie-Tooth (CMT) disease, one of the most common inherited neurological disorders. Abnormal development of myelin in the CNS results in disorders known as leukodystrophies. We previously described the severe peripheral neuropathy CMT4J, caused by mutation of the human FIG4/SAC3 gene encoding an evolutionarily conserved lipid phosphatase that regulates intracellular vesicle trafficking along the endo-lysosomal pathway. The main objectives of our research are to understand the molecular mechanisms by which FIG4 deficiency disrupts myelin formation, and to develop treatment strategies for CMT4J in a preclinical model. Mutant mice with global loss of Fig4 expression (Fig4-/-) exhibit dramatic reduction of myelin in the CNS and PNS, severe tremor, and juvenile lethality. Electrophysiological recordings revealed slowed conduction of electrical impulses in sciatic and optic nerves. Surprisingly, the myelin defects in Fig4-/- mice can be "rescued" by neuron-specific expression of wildtype Fig4. Based on these observations, we hypothesize that loss of Fig4 disrupts neuron-specific signaling mechanisms required for myelination. In Specific Aim 1 and Aim 2 we use a combination of mouse genetics and proteomics to identify the neuronal myelination signals that are disrupted in Fig4 mutant mice and to determine the temporal requirement for Fig4 in vivo. These experiments will provide new mechanistic insights into the neuronal signals that direct myelinogenesis. To model human CMT4J, we developed transgenic mice that ubiquitously express low levels of the human disease allele Fig4-I41T on a Fig4-/- background (CMT4J mice). These mice exhibit hypomyelination comparable to that of Fig4-/- mice, but survive to adulthood with many neurologic features of the human disease. Since we have shown that transgenic expression of Fig4 in neurons is sufficient to drive myelination, we propose a gene therapy study in Specific Aim 3. Dorsal root ganglion neurons (PNS) and retinal ganglion cells (CNS) of CMT4J mice will be transduced with viral vectors to express wildtype Fig4. Myelination, nodal structure, and nerve conduction velocity in sciatic or optic nerve will be monitored as indicators of efficacy. Restoration of myelination by Fig4 gene therapy in mice would demonstrate a new therapeutic avenue for patients suffering from myelination disorders. PUBLIC HEALTH RELEVANCE: Malformation, degeneration or acute damage to myelin in the nervous system is observed in a broad spectrum of white matter disorders including leukodystrophies and multiple sclerosis in the central nervous system and Charcot-Marie-Tooth disease in the peripheral nervous system. The research described in this project uses a novel mouse genetic model to probe the molecular and biochemical processes involved in white matter disease. A better understanding of these processes will lay the foundation for the development of treatment strategies for nervous system white matter disorders.