Stem Cell-Derived Motoneurons in the Adult mammalian CNS

描述(申请人提供)：我们发现运动神经元分化的ES细胞移植到运动神经元损伤的成年大鼠体内后存活，并且许多细胞表现出成熟运动神经元的免疫组织化学特征。在这个项目中，我们将确定促进在受损的成年哺乳动物中枢神经系统中产生功能性ES细胞衍生运动神经元的策略。在具体目标1中，我们将确定宿主应用和体外策略，以优化急性损伤脊髓中运动神经元的分化和存活。我们将确定这些细胞在几个分化阶段(即多能干细胞、承诺神经前体、承诺运动神经元前体或有丝分裂后运动神经元)的每个阶段存活和移植到大鼠脊髓内的能力。此外，我们还将探索细胞移植到的环境是否会调节生存和分化。在特定的目标2中，我们将测试增强ES细胞来源的轴突克服中枢神经系统白质介导的轴突排斥的能力的方法。我们将调节Nogo-R、P75-NTR和RhoA的活性，以便使移植的ES细胞在导航到白质周围时不太容易受到宿主排斥线索的影响。在具体目标3中，我们将测试增强ES细胞来源的运动神经元向远端延伸轴突并允许在体内形成功能性神经肌肉连接的方法。基于ES来源运动神经元上神经营养素受体的表达，我们建议在外周表达同源配体(包括IGF-1和GDNF)，以吸引ES细胞来源的运动轴突向远端生长并神经支配肌肉。逆行标记、电生理测试和行为学研究将确定这些策略在产生功能运动神经元和恢复瘫痪动物运动单位方面的有效性。这些研究将为ALS和SMA等运动神经元疾病以及脊髓损伤和其他脊柱疾病确定一种重要的移植方法。

DESCRIPTION (provided by applicant): We have found that motoneuron-differentiated ES cells survive when transplanted into adult rats with a motoneuron injury and that many of the cells exhibit immunohistochemical features of mature motoneurons. In this project, we will determine strategies that facilitate the generation of functional ES cell-derived motoneurons in the injured adult, mammalian CNS. In specific aim 1, we will define host-applied and ex vivo strategies that optimize motoneuron differentiation and survival in the acutely damaged spinal cord. We will determine the ability of these cells to survive and engraft within the rat spinal cord at each of several differentiation stages (i.e. pluripotent stem cell, committed neural precursor, committed motoneuron precursor or postmitotic motoneuron). Additionally, we will explore whether the environment into which the cells are transplanted modulates survival and differentiation. In specific aim 2, we will test approaches that enhance the ability of ES cell-derived axons to overcome CNS white matter-mediated axonal repulsion. We will modulate the activity of NOGO-R, P75-NTR and RhoA in order to transiently render the transplanted ES cells less susceptible to host repulsive cues as they navigate surrounding white matter. In specific aim 3, we will test approaches that enhance the ability of ES cell-derived motoneurons to extend axons distally and allow the formation of functional neuromuscular junctions in vivo. Based on expression of neurotrophin receptors on ES-derived motoneurons, we propose to peripherally express cognate ligands (including IGF-1 and GDNF) that will attract ES-cell derived motor axons to grow distally and to innervate muscle. Retrograde labeling, electrophysiologic testing and behavioral studies will determine the effectiveness of these strategies in generating functional motoneurons and restoring motor units in paralyzed animals. These studies will define an important transplantation approach for motoneuron disease such as ALS and SMA and for spinal cord injury and other spinal disorders.