MULTIPRONGED APPROACH TO PROMOTE FUNCTIONAL AXONAL REGENERATION AFTER SCI

多管齐下促进 SCI 后功能性轴突再生

• 批准号: 9123155
• 负责人: Veronica Jean Tom
• 金额: $ 4.97万
• 依托单位: DREXEL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9123155
• 关键词: Address; Adult; Affect; American; Astrocytes; Axon; Behavioral; Cerebral Ischemia; Chondroitin ABC Lyase; Cicatrix; Clinical; Complex; Data; Distal; Embryonic Nervous System; Environment; Excitatory Synapse; Exogenous Factors; Extracellular Matrix; Fiber; Glypican; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Health; Heparan Sulfate Proteoglycan; Histologic; In Vitro; Injury; Lead; Lesion; Mediating; Modeling; Motor; Natural regeneration; Nerve Degeneration; Nervous system structure; Neuraxis; Neurons; Pathway interactions; Peripheral Nerves; Persons; Physiological; Population; Property; Protein Biosynthesis; Public Health; Quality of life; Ras homolog enriched in brain; Recovery; Recovery of Function; Sensory; Site; Spinal Cord; Spinal cord injury; Synapses; Testing; Tissues; Translating; Work; axon growth; axon regeneration; clinical application; improved; improved functioning; in vivo Model; mTOR protein; neurotrophic factor; novel; novel strategies; novel therapeutics; postsynaptic; presynaptic; reinnervation; relating to nervous system; repaired; research study; response; synaptogenesis

DESCRIPTION (provided by applicant): While the mature central nervous system has an overall diminished capacity for regrowth compared to an embryonic nervous system, axonal regeneration of some populations is possible when they are provided with a potently growth-permissive environment, such as one conferred by a peripheral nerve graft (PNG). Still, axons that are capable of regenerating into the graft fail to grow beyond the PNG to reinnervate spinal cord, at least partly because of the presence of the inhibitory glial scar that is present at the graft-host interface. Digesting the inhibitory extracellular matrix within the scar with chondroitinase ABC (ChABC) allows some axons to traverse the distal interface, synapse upon distal neurons, and mediate some behavioral recovery. Nonetheless, most of the axons remain in the graft. Thus it is important to develop strategies that enhance the ability of axons to regenerate out of a graft in order to increase their potential to improve function. We have preliminary data indicating that expressing constitutively active Rheb (caRheb) - a GTPase that is a critical regulatory component of protein synthesis - in adult neurons results in robust axonal outgrowth in an in vitro glial scar model and in vivo after SCI. In Aim 1, we will test the hypothesis that expressing caRheb after SCI will enhance regeneration into and beyond a PNG. We will use histological, physiological, and behavioral analyses to examine if caRheb augments axonal regeneration - especially following ChABC treatment of the distal graft interface - and if these regenerated axons form functional synapses that facilitate behavioral recovery. Any regeneration is irrelevant if these axons do not integrate into circuits. Though ChABC enhances plasticity, ChABC-induced sprouted/regenerated fibers do not always form synapses to impact function. Thus, it is also important to devise strategies to improve integration of these axons (i.. synaptogenesis) in order to exploit regrowth that we encourage. We recently demonstrated the proof-of-principle that providing a single exogenous factor enhances the integration of axons that regenerate out of a PNG. Recent work indicates that secreted glypican is sufficient to promote functional, excitatory synapse formation in vitro and behavioral recovery in a cerebral ischemia model in vivo. In Aim 2, we will test the hypothesis that providing exogenous glypican will enhance synaptogenesis of regenerated axons following SCI. We will graft a PNG into a SCI site that has been treated with ChABC to promote axonal regeneration. We will determine if infusing exogenous glypican into tissue surrounding the SCI site enhances synaptogenesis of these regenerated axons upon neurons distal to the PNG. We will gauge improvements in functional synapse formation physiologically and histologically by examining the colocalization of pre- and postsynaptic markers. We will also assess if enhanced synapse formation correlates with behavioral recovery. Because the strategies to be tested in Aims 1 and 2 take different approaches - enhancing the growth response and synaptogenesis, respectively - to promote functional repair after SCI, it is possible that even if the results in the previous Aims are incremental, combining the two approaches will lead to greater recovery than either alone. In Aim 3, we will use a novel, multipronged strategy to fill the lesion cavity with a growth-promoting substrate (PNG), enhance the growth response (caRheb), modulate the inhibitory properties of the glial scar (ChABC) and promote synaptogenesis (glypican). We will use histological and physiological analyses to examine if glypican enhances the functional integration of axonal regeneration beyond a ChABC-treated PNG interface that is induced by expressing caRheb and if this results in more robust behavioral recovery than what occurs with either caRheb or glypican treatment separately. These data will identify new therapeutic strategies that may eventually be translated to clinical use to improve the quality of life of persons with SCI.

描述（由申请人提供）：虽然与胚胎神经系统相比，成熟的中枢神经系统的再生能力总体减弱，但当为某些群体提供有效的生长许可环境（例如赋予的环境）时，它们的轴突再生是可能的。通过周围神经移植（PNG）。尽管如此，能够再生到移植物中的轴突无法生长到PNG之外以重新支配脊髓，至少部分是因为移植物-宿主界面处存在抑制性神经胶质疤痕。用软骨素酶 ABC (ChABC) 消化疤痕内的抑制性细胞外基质，使一些轴突能够穿过远端界面，在远端神经元上形成突触，并介导一些行为恢复。尽管如此，大多数轴突仍保留在移植物中。因此，重要的是制定增强轴突从移植物中再生的能力的策略，以增加其改善功能的潜力。我们的初步数据表明，在成年神经元中表达组成型活性的 Rheb (caRheb)（一种 GTP 酶，是蛋白质合成的关键调节成分）会产生强大的轴突。 体外神经胶质疤痕模型和 SCI 后体内的生长。在目标 1 中，我们将测试以下假设：SCI 后表达 caRheb 将增强 PNG 内和之外的再生。我们将使用组织学、生理学和行为分析来检查 caRheb 是否会增强轴突再生（尤其是在对远端移植物界面进行 ChABC 治疗后），以及这些再生的轴突是否形成促进行为恢复的功能性突触。如果这些轴突不整合到电路中，任何再生都是无关紧要的。尽管 ChABC 增强了可塑性，但 ChABC 诱导的发芽/再生纤维并不总是形成突触来影响功能。因此，制定策略来改善这些轴突的整合（即突触发生）以利用我们鼓励的再生也很重要。我们最近证明了提供单一外源因子可以增强从 PNG 再生的轴突整合的原理验证。最近的研究表明，分泌的磷脂酰肌醇蛋白聚糖足以促进体外功能性、兴奋性突触的形成和体内脑缺血模型的行为恢复。在目标 2 中，我们将检验以下假设：提供外源磷脂酰肌醇蛋白聚糖将增强 SCI 后再生轴突的突触发生。我们将把 PNG 移植到经过 ChABC 处理的 SCI 部位，以促进轴突再生。我们将确定将外源性磷脂酰肌醇蛋白聚糖注入 SCI 部位周围的组织中是否会增强这些再生轴突在 PNG 远端神经元上的突触发生。我们将通过检查突触前和突触后标记的共定位来衡量功能性突触形成在生理学和组织学上的改善。我们还将评估突触形成的增强是否与行为恢复相关。由于目标 1 和目标 2 中要测试的策略采用不同的方法（分别增强生长反应和突触发生）来促进 SCI 后的功能修复，因此即使前面的目标中的结果是增量的，也有可能将两种方法结合起来将比单独使用任何一种方法带来更好的恢复。在目标 3 中，我们将使用一种新颖的、多管齐下的策略，用促生长物质填充病变空腔。 底物 (PNG)、增强生长反应 (caRheb)、调节神经胶质疤痕 (ChABC) 的抑制特性并促进突触发生 (磷脂酰肌醇蛋白聚糖)。我们将使用组织学和生理学分析来检查磷脂酰肌醇蛋白聚糖是否增强了轴突再生的功能整合，超出了通过表达 caRheb 诱导的 ChABC 处理的 PNG 界面，以及这是否会导致比单独使用 caRheb 或磷脂酰肌醇蛋白聚糖治疗更稳健的行为恢复。这些数据将确定新的治疗策略，最终可能转化为临床应用，以改善 SCI 患者的生活质量。