一种通过重建皮质脊髓神经支配恢复脑卒中后偏瘫肢体运动功能的创新性策略

Stroke is the leading cause of serious, long-term disability in adults. Unfortunately, there are no efficacious therapies available for the vast majority of chronic stroke patients. Since the spinal motoneurons are the final common pathway of brain-controlled movements of the peripheral muscles, we hypothesize that re-establishment of corticospinal innervation in the spinal gray matter provides a physical substrate for functional recovery after stroke. Focusing on the denervated spinal motoneurons which innervate the stroke-impaired muscles, we propose a novel peripheral approach, to over-express brain-derived neurotrophic factor (BDNF) in the spinal motoneurons via recombinant adeno-associated virus (AAV)-BDNF intramuscular injection，and over-express its receptor TrkB in the cortical neurons and the corticospinal tract (CST) axons by intranasal AAV-TrkB delivery. BDNF binding to TrkB stimulates CST axonal sprouting, outgrowth and synapse formation in the denervated side of the spinal gray matter, and thereby promotes motor functional recovery after stroke. We will employ a middle cerebral artery occlusion (MCAo) model in adult rats, combined with behavioral, neuroanatomical, and electrophysiological technique to investigate the therapeutic effect of intramuscular AAV-BDNF and intranasal AAV-TrkB treatment promoting neurological recovery after stroke. To demonstrate the therapeutic effect of AAV-BDNF and AAV-TrkB administration on CST axonal remodeling to promote motor functional recovery after stroke, we will perform a battery of behavioral tests to evaluate neurological outcome after stroke. Electromyograms (EMG) will be recorded from the stroke-impaired forelimb muscles in response to intracortical microstimulations (ICMS) on bilateral motor cortices to assess the functional re-establishment of corticospinal innervation after stroke. The neural pathways between the motor cortices and spinal cord will be traced by biotin-conjugated dextran amine (BDA) and rhodamine-conjugated dextran amine (RDA) anterogradely, and with trans-synaptic pseudorabies virus (PRV) retrogradely. Synaptic remodeling in the denervated side of the spinal cord will be assessed with immunohistological staining and transmission electron microscopy. To determine the contribution of CST subsets to neurological recovery, rats will be subjected to MCAo followed by unilateral or bilateral pyramidotomy. Our studies may greatly impact the development of novel and efficacious treatments for a large population of stroke patients with a significant unmet medical need of efficacious stroke treatment beyond the acute stage.

由于脊髓运动神经元是大脑控制的外周肌肉运动的最终公共通路，我们推测皮质脊髓神经支配的重建乃中风后功能恢复的物质基础。针对支配中风偏瘫肌肉的脊髓运动神经元, 我们提出了一种新的外周途径，通过在偏瘫肌肉中注射重组腺相关病毒（AAV）增加脑源性神经营养因子（BDNF）在脊髓运动神经元中的表达。同时经鼻给与AAV载体增加其受体TrkB表达水平。BDNF与TrkB结合将刺激皮质脊髓束（CST）轴突朝向这些失神经支配的脊髓运动神经元发芽、生长，并与之形成突触联结，从而恢复中风后偏瘫肢体的运动功能。我们将在成年大鼠中采用大脑中动脉闭塞（MCAo）模型，结合行为学，神经解剖学和电生理学技术，来研究肌内注射AAV-BDNF及经鼻给与AAV-TrkB促进中风后运动功能恢复的治疗效果。本研究的成果，可望为开发临床迫切需要的亚急性及慢性期卒中及其他神经损伤患者创新性有效治疗方法带来突破性进展。

脑卒中导致幸存者长期残疾。BDNF/TrkB 在中枢神经系统 (CNS) 的突触可塑性和突触传递中发挥重要作用，促进神经功能恢复。在这项研究中，我们进行了非侵入性治疗方法，使用携带 BDNF 或 TrkB 编码基因的腺相关病毒 (AAV) 载体肌​​肉注射到中风受伤的前肢肌肉中，或进行鼻内给药。在永久性大鼠大脑中动脉闭塞 (MCAO) 模型中，我们评估了 AAV-BDNF 和 AAV-TrkB 联合治疗对皮质脊髓连接的运动功能恢复和突触可塑性的影响。我们的结果表明，BDNF 或 TrkB 基因在脊髓前角神经元和大脑皮层神经元中转导。与单独使用 AAV 载体治疗相比，行为和电生理学结果表明，联合治疗显着改善了卒中后上肢运动功能恢复和神经传递效率。BDA 示踪、免疫荧光染色、qRT-PCR 和突触超微结构的透射电镜结果表明，联合治疗不仅有效增加突触蛋白 I、PSD-95 和 GAP-43 的表达，而且促进轴突重塑和恢复异常突触结构。这些发现提供了一种增强神经可塑性的新策略和临床治疗脑卒中的潜在手段。