DEATH VS GROWTH PROGRAMME IN AXONAL INJURED MOTORNEURONS

轴突损伤运动神经元的死亡与生长计划

• 批准号: 2272939
• 负责人: WUTIAN WU
• 金额: $ 9.22万
• 依托单位: EASTERN VIRGINIA MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-04-01 至 2000-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2272939
• 关键词: axon reaction; calcitonin gene related peptide; cell cycle proteins; denervation; gene induction /repression; growth factor receptors; in situ hybridization; laboratory rat; motor neurons; nervous system regeneration; nervous system transplantation; neuronal transport; neurotrophic factors; nitric oxide synthase; regulatory gene; spinal nerves

Recent investigations have shown that injured neurons express a programme, presumably involving the sequential switching of genes, that may control regrowth and axonal regeneration or induces cell death. The precise mechanisms and roles of such gene switching are poorly understood. Preliminary data from the PI's laboratory have demonstrated that spinal motoneurons in adult rat underwent a series of gene expression changes following axonal injuries. Some of these gene expressions seem to be related to regenerative processes, others seem to be associated with neuronal death. Peripheral nerve implantation seems to alter some gene expressions and reduce the death of motoneurons due to axonal injuries. The long-term goal of the proposed project is to study the mechanism underlying neuronal degeneration and regeneration, and to find new approaches which could prevent degeneration, promote regeneration, and improve functional recovery after neuronal injury. Four specific aims are included in the present proposal. (1). To examine gene expression changes in spinal motoneurons in response to axonal injuries in adult animals. Emphases will be focused on immediate early genes, neurotrophin receptor genes, growth associated genes, and nitric oxide synthase. (2). To investigate whether gene expression changes are associated with neuronal degeneration or regeneration. If so, to determine which is related to degeneration and which is related to regeneration. (3). To test the effect of a microsurgical procedure, the peripheral nerve system (PNS) graft implantation, on motoneuron survival and regeneration following axonal injury, and to determine which gene expression is associated with the survival and regeneration of injured motoneurons. (4). To investigate whether neurotrophic factors could prevent motoneuron death and promote regeneration following axonal injury. If so, to determine whether such effect is achieved by modifying certain gene expressions in injured motoneurons. In the proposed experiments, adult motoneurons will be used as experimental model. Advantage of this model is that neuronal death or regeneration in spinal motoneurons can be reliably manipulated by experimental procedures. In the proposed experiments, gene expression changes in spinal motoneurons will be studied under three different injury conditions which include: 1). distal spinal root axotomy (which does not cause motoneuron death), 2). spinal root avulsion (which causes a significant motoneuron death), and 3). root avulsion followed by PNS graft implantation or/and neurotrophic factor treatments (which prevents injured motoneurons from death and allows them to regenerate). These experimental procedures allow us to examine gene expressions and their modifications in different cases of motoneurons after axonal injuries. Gene expressions will be examined using immunocytochemistry and in situ hybridization techniques. Cell death will be assessed by neuronal counts following the experimental manipulations. Axonal regeneration will be assessed by a double fluorescent retrograde labeling technique: Data obtained from these experiments should provide a better understanding of events leading to neuronal death and plasticity and may lead to therapeutic interventions that prevent neuronal death and enhance regeneration.

最近的调查表明，受伤的神经元表达了一个程序， 大概涉及基因的顺序切换，可能控制 再生和轴突再生或诱导细胞死亡。 确切的 这种基因转换的机制和作用知之甚少。 PI实验室的初步数据证明了脊柱 成年大鼠的运动神经元发生了一系列基因表达变化 轴突损伤后。 这些基因表达中的一些似乎是 与再生过程有关，其他似乎与 神经元死亡。 周围神经植入似乎改变了某些基因 表达并减少由于轴突损伤引起的运动神经元的死亡。 拟议项目的长期目标是研究机制 潜在的神经元变性和再生，并找到新的 可以防止退化，促进再生的方法 改善神经元损伤后的功能恢复。 四个具体目标是 包括在本提案中。 （1）。 检查基因表达变化 在脊柱运动神经元中，响应成年动物的轴突损伤。 重点将集中于立即的早期基因，神经营养蛋白受体 基因，相关基因和一氧化氮合酶。 （2）。 到 研究基因表达变化是否与神经元有关 退化或再生。 如果是这样，以确定哪个与 变性，与再生有关。 （3）。 测试 显微外科手术，外围神经系统（PNS）的影响 移植物植入，运动神经元的生存和再生之后 轴突损伤，并确定哪种基因表达与 受伤的运动神经元的生存和再生。 （4）。 调查 神经营养因素是否可以预防运动神经元死亡并促进 轴突损伤后再生。 如果是这样，以确定是否 通过修改受伤的某些基因表达来实现效果 运动神经元。 在拟议的实验中，成年运动神经元将被用作实验 模型。 该模型的优势是神经元死亡或再生 可以通过实验程序可靠地操纵脊柱运动神经元。 在拟议的实验中，基因表达在脊柱运动神经元中的变化 将在三种不同的伤害条件下进行研究：1）。 脊柱远端根轴切开术（不会导致运动神经元死亡），2）。 脊柱根撕脱（会导致明显的运动神经元死亡），而3）。 根撕裂，然后是PNS移植植入或/和神经营养因子 治疗（阻止受伤的运动神经元死亡并允许他们 再生）。 这些实验程序使我们能够检查基因 表达式及其在不同病例的运动神经元之后进行的修改 轴突受伤。 基因表达将使用 免疫细胞化学和原位杂交技术。 细胞死亡 在实验操作后通过神经元计数评估。 轴突再生将通过双荧光逆行评估 标签技术： 从这些实验获得的数据应提供更好的理解 导致神经元死亡和可塑性的事件，可能导致 预防神经元死亡并增强的治疗干预措施 再生。