Neuronal regulation of myelination

描述（由申请人提供）：在脊椎动物（包括人类）中，外周（PNS）和中枢神经系统（CNS）中的快速神经元通讯依赖于适当的髓鞘形成。PNS中的髓鞘形成细胞是雪旺细胞（SC），CNS中的髓鞘形成细胞是少突胶质细胞（OL）。这些特化的细胞包裹神经突起，从而促进电脉冲的快速传播。PNS髓磷脂在几种类型的腓骨肌萎缩症（CMT）中有缺陷，这是最常见的遗传性神经系统疾病之一。中枢神经系统中髓鞘的异常发育导致称为脑白质营养不良的疾病。我们先前描述了严重的周围神经病变CMT 4J，由编码进化上保守的脂质磷酸酶的人FIG 4/SAC 3基因突变引起，所述脂质磷酸酶调节胞内囊泡沿内-溶酶体途径的运输沿着。我们研究的主要目的是了解FIG 4缺陷破坏髓鞘形成的分子机制，并在临床前模型中开发CMT 4J的治疗策略。具有Fig 4表达的整体缺失的突变小鼠（图4-/-）表现出CNS和PNS中髓磷脂的显著减少、严重震颤和幼年致死性。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，图4-/-小鼠中的髓鞘缺陷可以通过野生型的神经元特异性表达来“拯救”图4。基于这些观察，我们假设Fig 4的缺失破坏了髓鞘形成所需的神经元特异性信号传导机制。在特定目标1和目标2中，我们使用小鼠遗传学和蛋白质组学的组合来鉴定在图4突变小鼠中被破坏的神经元髓鞘形成信号，并确定图4在体内的时间要求。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CMT 4J，我们开发了在Fig 4-/-背景上普遍表达低水平的人疾病等位基因Fig 4-I41 T的转基因小鼠（CMT 4J小鼠）。这些小鼠表现出与图4-/-小鼠相当的髓鞘形成不足，但存活至成年，具有人类疾病的许多神经学特征。由于我们已经表明图4在神经元中的转基因表达足以驱动髓鞘形成，因此我们在特定目标3中提出了基因治疗研究。CMT 4J小鼠的背根神经节神经元（PNS）和视网膜神经节细胞（CNS）将用病毒载体转导以表达野生型图4。将监测坐骨神经或视神经中的髓鞘形成、结节结构和神经传导速度，作为疗效指标。通过图4基因疗法在小鼠中恢复髓鞘形成将证明患有髓鞘形成障碍的患者的新治疗途径。

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.