Functional Analysis of Myelin Inhibitors in Spinal Axon Regeneration Failure

描述(申请人提供)：成年哺乳动物中枢神经系统(CMS)的轴突再生失败至少部分归因于中枢神经系统髓鞘的抑制性质。Nogo、髓鞘相关糖蛋白(MAG)和少突胶质细胞髓磷脂糖蛋白(OMGp)是三种经典的髓鞘来源的神经突起生长抑制剂，它们对体外培养的神经突起生长具有很强的抑制作用。然而，核心问题仍然是这些抑制物在体内中枢神经系统轴突再生失败中的作用。我们的中心假设是髓鞘来源的轴突生长抑制剂Nogo、MAG和OMgp在阻止脊髓轴突再生方面发挥着重要的和潜在的多余作用。总的方法是慢性或急性地检查一种或多种髓鞘抑制剂基因缺失的小鼠脊髓中的轴突再生，具体解决发育补偿和功能冗余的问题。通过急性基因缺失，我们解决了生殖系突变的发育补偿问题。通过同时删除这三个抑制物，我们解决了功能冗余的问题。目的1.通过检测急性缺失Nogo或OMgp对皮质脊髓和延髓5-羟色胺能纤维再生的影响，探讨Nogo和OMgp在再生失败中的作用。我们将使用一种可诱导的基因敲除系统来精确删除少突胶质细胞中的Nogo或OMgp，然后检测皮质脊髓束(CST)和纹状体5-羟色胺能纤维束的再生反应。目的2.通过对缺乏Nogo、MAG和OMgp的小鼠皮质脊髓和延髓5-羟色胺能纤维束的再生潜能的研究，探讨Nogo、MAG和OMgp在中枢神经系统轴突再生失败中的联合作用。目的3.检测提高脊髓固有生长潜能与去除髓鞘抑制物在促进脊髓轴突再生方面是否具有协同作用。先前的研究表明，损毁背根神经节(DRG)神经元的外周分支可以增强神经元的内在生长潜力，促进体内中心支的再生。我们将测试这种增强的再生在Nogo/MAG/OMgp三重突变体中是否进一步增强，以及删除这三种髓鞘抑制物与条件性损伤之间是否存在协同效应。总之，这些研究将为髓鞘抑制剂在脊髓轴突再生失败中的作用提供重要的见解。了解这些髓鞘抑制物在中枢神经系统轴突再生中的作用，对于设计任何针对这一组分子的促进轴突再生和脊髓修复的治疗干预措施至关重要。

DESCRIPTION (provided by applicant): Axon regeneration failure in the adult mammalian central nervous system (CMS) has been attributed at least in part to the inhibitory nature of the CNS myelin. Three "classical" myelin-derived neurite outgrowth inhibitors, Nogo, myelin-associated glycoprotein (MAG) and oligodendrocyte-myelin glycoprotein (OMgp), have been identified that exhibit potent inhibitory activity on neurite outgrowth in vitro. However, the central question remains as to the contribution of these inhibitors to CNS axon regeneration failure in vivo. Our central hypothesis is that myelin-derived neurite growth inhibitors Nogo, MAG and OMgp play a significant and potentially redundant role in blocking spinal axon regeneration. The overall approach is to examine axon regeneration in the spinal cord of mice with gene deletion in one or more myelin inhibitors chronically or acutely, specifically addressing the issues of developmental compensation and functional redundancy. By acute gene deletion, we address the issue of developmental compensation in germline mutants. By deleting the three inhibitors simultaneously, we address the issue of functional redundancy. Aim 1. To assess the role of Nogo and OMgp in regeneration failure by examining the effect of acutely deleting Nogo or OMgp on corticospinal and raphespinal serotonergic fiber regeneration. We will employ an inducible knockout system to acutely delete Nogo or OMgp in oligodendrocytes and then examine the regenerative response of the corticospinal tract (CST) and the raphespinal serotonergic fiber tract. Aim 2. To assess the combined contribution of Nogo, MAG and OMgp in CNS axon regeneration failure by ascertaining the regeneration potential of the corticospinal and raphespinal serotonergic fiber tracts in mice deficient in all three inhibitors. Aim 3. To test whether increasing the intrinsic growth potential has a synergistic effect with removing myelin inhibitors in promoting spinal axon regeneration. Previous studies indicate that a conditioning lesion to the peripheral branch of the dorsal root ganglion (DRG) neurons augments the intrinsic growth potential of the neurons and promotes the regeneration of the central branch in vivo. We will test whether this enhanced regeneration is further enhanced in Nogo/MAG/OMgp triple mutant, and whether there is a synergistic effect between deleting the three myelin inhibitors and a conditioning lesion. Together, these studies will provide important insight into the role of myelin inhibitors in spinal axon regeneration failure. Understanding the role of these myelin inhibitors in CNS axon regeneration is crucial to the design of any therapeutic intervention to promote axon regeneration and spinal cord repair targeting this group of molecules.