Neurotransplantation and Training to Promote Recovery of Chronic SCI Cats

神经移植和训练促进慢性脊髓损伤猫的康复

• 批准号: 8258144
• 负责人: John D. Houle
• 金额: $ 37.73万
• 依托单位: DREXEL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8258144
• 关键词: Acute; Address; Adult; Affect; Animal Model; Animals; Axon; Back; Behavioral; Biomechanics; Brain; Chemicals; Chest; Chondroitin Sulfate Proteoglycan; Chondroitinases; Chronic; Cicatrix; Combined Modality Therapy; Communication; Data; Detection; Distal; Electric Stimulation; Environment; FOS gene; Felis catus; Financial compensation; Forelimb; Fostering; Future; Gait; Growth; Hindlimb; Human; Hyaluronidase; Implant; Individual; Injury; Interneurons; Investigation; Joints; Label; Learning; Lesion; Limb structure; Link; Locomotion; Locomotor Recovery; Measures; Modeling; Muscle; Natural regeneration; Nerve; Neurons; Neurorehabilitation; Operative Surgical Procedures; Organ Transplantation; Pain; Pathway interactions; Patients; Perception; Peripheral Nerves; Peripheral nerve injury; Physical therapy; Physiological; Procedures; Process; Rattus; Recovery; Recovery of Function; Rehabilitation therapy; Research; Risk; Role; Sensory Ganglia; Site; Source; Spinal; Spinal Cord; Spinal Injuries; Spinal cord injury; Synapses; Techniques; Testing; Thromboplastin; Time; Tissues; Training; Translations; Transplantation; Work; axon growth; axon regeneration; base; clinical application; clinically relevant; design; immunocytochemistry; immunoreactivity; improved; injured; kinematics; locomotor tasks; neurotrophic factor; novel; pre-clinical; reconstruction; regenerative; repaired; research study; response; scale up; success; synaptogenesis; treatment strategy

DESCRIPTION (provided by applicant): Peripheral nerve grafts (PNGs) provide an excellent substratum for axonal regrowth; they can direct regenerating axons towards a specific target and they facilitate electrophysiological experimentation to detect synaptic connectivity between regenerating axons and distal spinal cord neurons. A major impediment to this and all other transplantation approaches after spinal cord injury is the poor growth of axons out of the graft back into the host spinal cord. We have combined Chondroitinase (ChABC, to digest inhibitory chondroitin sulfate proteoglycan molecules) with PN grafting in rats and have anatomical and electrophysiological evidence for functional synapse formation by injured, regenerating axons in both acute and delayed (chronic injury) treatment paradigms. Recently we replicated this rat acute PNG approach in cats where we observed thousands of axons regenerating into the graft, a small percentage of which extended from the graft into the spinal cord distal to the injury, and spinal neurons synaptically activated (determined by c-Fos immunoreactivity) after electrical stimulation of the nerve graft. While we will continue to use rat models for expanding our treatment repertoire, the objective of this study is to focus on application of our treatment strategies to chronically injured cats as a necessary preclinical step before translation into human research. The cat model permits us to investigate issues related to the scaling up of a transplantation model, cats are easily trained to perform locomotor tasks, and recovery of function can be assessed by kinematic and electrophysiological measures. The biomechanics of locomotion are better defined in cats and cats have a hindlimb gait that is close to human than is the rat. The proposed work also will provide information about the ability to effectively treat glial scarring in a large animal, the ability to promote structural and functional regeneration in a large animal with a chronic injury and the potential for rehabilitation training to foster regeneration and functional recovery. There are 2 Specific Aims for this project. 1) We will identify the source and extent of axonal regeneration into a PNG after chronic injury and test whether these axons form functional connections across the lesion. 2) We will test whether the start time of physical rehabilitation affects outgrowth, integration and/or synaptic activity of regenerating axons. A combination of treatment strategies will be used, including transplantation, ChABC treatments and treadmill training to promote activity dependent plasticity. Structural repair will be assessed by anatomical tract tracing and immunocytochemical labeling; forelimb-hindlimb coordination will be assessed by kinematic and electromyogram (EMG) analysis; functional reconnection will be measured during electrophysiological stimulation of the graft and by c-fos expression in synaptically activated neurons. Surgical intervention after SCI usually is not an option until the patient is stabilized, thus the majority of individuals with SCI likely will be chronically injured before a treatment strategy for repair is initiated. Our work with chronically injured rats demonstrates the ability to promote long distance regeneration with formation of functionally active synapses distal to an injury. The proposed study will take advantage of the treatment approaches that have been (and are being) developed with chronically injured rats, but will apply them to a large animal model of SCI. This preclinical advancement is a crucial step towards translation to a clinical application. We propose a unique approach to address a very important aspect of SCI, i.e. chronic injury in a large animal model. Locomotor training of injured cats has been carried out by numerous labs, but not in a situation where axon regeneration is facilitated. This will be a novel application of neuroregeneration and neurorehabilitation techniques to increase our understanding of the potential for repair after SCI. PUBLIC HEALTH RELEVANCE: Different transplantation models have been used to demonstrate that under the right conditions neurons in spinal cord injured adult rats will regenerate their nerve processes (axons) into and through the transplant, forming a bridge across the lesion to restore lines of communication between the brain and spinal cord. Part of the success that has been demonstrated with these models involves additional treatment of the spinal cord tissue adjacent to the injury, to either increase the presence of growth promoting molecules or to decrease the presence of growth inhibitory molecules. An important issue to address concerns the time after injury when a treatment might be most effective and we have explored this question by delaying treatment for weeks to months after injury. We have demonstrated that chronically injured neurons in rats retain the capacity for regeneration for long (at least 12 months) post injury periods. This observation directly impacts the overwhelming number of spinal cord injured patients because of the perception that most surgical interventions should be delayed until the individual is stable and opportunities for spontaneous recovery have subsided. As a prelude to work with human patients we have pursued our peripheral nerve graft studies in a larger animal model, spinal cord injured cats. The purpose of this study was to determine if multiple surgical procedures could be performed on the cat spinal cord without causing significant pain or discomfort to the animal and if peripheral nerve grafts used to bridge the lesion in rats would be equally effective in cats, providing a channel for regenerating axons to form functional synaptic contacts with spinal cord neurons. We have data demonstrating the successful application of our combination of treatments to the adult cat spinal cord when carried out immediately after injury. The present proposal will extend this observation to the chronically injured cat model where treatment strategies will be delayed for several months, with the objective of advancing the use of a combination of treatment strategies towards translation into human application. The proposed work will provide valuable information about the ability to perform surgical reconstruction of spinal cord circuitry in a large animal, to determine if size might be a hindrance to successful regeneration. It also will provide an understanding of the potential to promote structural and functional recovery in a large animal with a longstanding (chronic) injury and of the ability to foster greater recovery through aggressive physical rehabilitation training. We feel that results of this study would have direct clinical relevance with the major risk being in not pursuing this line of investigation.

描述（由申请人提供）：周围神经移植物（PNG）为轴突再生提供了极好的基质；它们可以将再生轴突导向特定目标，并促进电生理学实验以检测再生轴突和远端脊髓神经元之间的突触连接。脊髓损伤后这种移植方法和所有其他移植方法的一个主要障碍是从移植物返回宿主脊髓的轴突生长不良。我们将软骨素酶（ChABC，用于消化抑制性硫酸软骨素蛋白聚糖分子）与大鼠 PN 移植相结合，并获得了急性和延迟（慢性损伤）治疗范例中受损、再生轴突形成功能性突触的解剖学和电生理学证据。最近，我们在猫身上复制了这种大鼠急性 PNG 方法，观察到数千个轴突再生到移植物中，其中一小部分从移植物延伸到损伤远端的脊髓，并且脊髓神经元突触激活（由 c-Fos 确定）神经移植物电刺激后的免疫反应性）。虽然我们将继续使用大鼠模型来扩展我们的治疗方案，但本研究的目的是重点将我们的治疗策略应用于慢性受伤的猫，作为转化为人类研究之前的必要临床前步骤。猫模型使我们能够研究与移植模型放大相关的问题，猫很容易被训练来执行运动任务，并且可以通过运动学和电生理学测量来评估功能的恢复。猫的运动生物力学得到了更好的定义，并且猫的后肢步态比老鼠更接近人类。拟议的工作还将提供有关有效治疗大型动物神经胶质疤痕的能力、促进慢性损伤大型动物结构和功能再生的能力以及促进再生和功能恢复的康复训练潜力的信息。该项目有 2 个具体目标。 1）我们将确定慢性损伤后 PNG 中轴突再生的来源和程度，并测试这些轴突是否在病变中形成功能连接。 2）我们将测试身体康复的开始时间是否影响再生轴突的生长、整合和/或突触活动。将结合使用治疗策略，包括移植、ChABC 治疗和跑步机训练，以促进活动依赖性可塑性。将通过解剖道追踪和免疫细胞化学标记来评估结构修复；将通过运动学和肌电图（EMG）分析来评估前肢-后肢协调性；功能重新连接将在移植物的电生理刺激过程中以及通过突触激活的神经元中的 c-fos 表达来测量。在患者病情稳定之前，SCI 后通常不能选择手术干预，因此大多数 SCI 患者在开始修复治疗策略之前可能会受到慢性损伤。我们对慢性损伤大鼠的研究表明，通过在损伤远端形成功能活跃的突触来促进长距离再生的能力。拟议的研究将利用已经（和正在）针对慢性损伤大鼠开发的治疗方法，并将其应用于大型 SCI 动物模型。这一临床前进展是转化为临床应用的关键一步。我们提出了一种独特的方法来解决 SCI 的一个非常重要的方面，即大型动物模型中的慢性损伤。许多实验室已经对受伤的猫进行了运动训练，但没有在促进轴突再生的情况下进行。这将是神经再生和神经康复技术的一个新应用，以增加我们对 SCI 后修复潜力的了解。 公共健康相关性：不同的移植模型已被用来证明，在适当的条件下，脊髓损伤的成年大鼠的神经元将在移植物中并通过移植物再生其神经突（轴突），形成跨越病变的桥梁，以恢复之间的通讯线路。大脑和脊髓。这些模型所证明的部分成功涉及对损伤附近的脊髓组织进行额外治疗，以增加生长促进分子的存在或减少生长抑制分子的存在。需要解决的一个重要问题是受伤后治疗可能最有效的时间，我们通过在受伤后推迟数周至数月的治疗来探讨这个问题。我们已经证明，大鼠中慢性损伤的神经元在损伤后很长一段时间（至少 12 个月）仍保留再生能力。这一观察结果直接影响了绝大多数脊髓损伤患者，因为人们认为大多数手术干预措施应推迟到个体稳定且自然恢复的机会减少为止。作为与人类患者合作的前奏，我们在更大的动物模型（脊髓损伤的猫）中进行了周围神经移植研究。本研究的目的是确定是否可以对猫脊髓进行多次外科手术而不会对动物造成明显的疼痛或不适，以及用于桥接大鼠病变的周围神经移植物是否对猫同样有效，从而提供再生轴突与脊髓神经元形成功能性突触接触的通道。我们有数据证明，在受伤后立即对成年猫脊髓进行联合治疗是成功的。目前的提案将把这一观察扩展到慢性受伤的猫模型，其中治疗策略将被推迟几个月，目的是推进治疗策略组合的使用，以转化为人类应用。拟议的工作将提供有关在大型动物中进行脊髓回路手术重建的能力的有价值的信息，以确定尺寸是否可能成为成功再生的障碍。它还将使人们了解促进长期（慢性）损伤的大型动物的结构和功能恢复的潜力，以及通过积极的身体康复训练促进更大程度恢复的能力。我们认为这项研究的结果将具有直接的临床相关性，主要风险是不进行这一研究。