Overcoming glial scar inhibitions on axonal growth

描述(申请人提供)：中枢神经系统轴突损伤后，促进神经功能障碍恢复的药物治疗极其有限。不允许轴突生长的环境至少部分导致了成年中枢神经系统的生长失败。具体地说，几组抑制分子强烈抑制中枢神经系统损伤后的轴突延伸，包括由胶质瘢痕产生的硫酸软骨素蛋白多糖(CSPGs)。CSPG是胶质瘢痕的主要抑制成分，是轴突再生的主要障碍。尽管已有几种策略被报道，但局部应用细菌软骨素酶ABC消化CSPGs是克服中枢神经系统损伤后CSPGs生长抑制的主要体内方法。然而，重要的缺点阻碍了这种酶作为轴突损伤患者的治疗选择，包括不能完全去除CSPG中的抑制成分，在体温下酶活性时间短，以及无法穿越血脑屏障。在这项建议中，我们的目标是开发新的治疗中枢神经系统轴突损伤的策略，基于对CSPGs的单独抑制或与我们先前确定的方法相结合。我们假设CSPGs的多肽拮抗剂将促进中枢神经系统损伤小鼠的形态和功能恢复。利用生物信息学的方法定义了几个CSPG的保守元件，我们已经确定了两个CSPG的选择性多肽拮抗剂。我们的初步研究表明，这些低纳摩尔浓度的多肽主要克服了神经元培养中CSPGs的轴突生长限制。全身应用CSPG阻滞肽显著改善体内中枢神经系统轴突损伤小鼠的行为恢复。在这项研究中，我们将表征这些CSPG拮抗肽在小鼠脊髓损伤(SCI)模型中的治疗潜力。除CSPG外，许多抑制分子通过激活收敛的RhoA或糖原合成酶激酶32(GSK-32)在细胞内参与抑制轴突生长。最近，我们已经证明用布洛芬灭活RhoA或用锂灭活GSK-32可以克服不同分子对RhoA的生长抑制，并显著促进脊髓损伤啮齿动物下行运动神经元的轴突生长和运动恢复。因此，我们的目标也是通过将CSPG阻断肽与抑制RhoA的布洛芬或GSK-32灭活锂结合起来，刺激脊髓损伤小鼠更戏剧性的轴突再生，这两种药物在人类中广泛使用。使用我们的新型CSPG拮抗剂，单独使用或与布洛芬或锂联合使用，可以通过促进轴突再生和功能恢复，显著提高我们治疗成年哺乳动物中枢神经系统轴突损伤的能力。 公共卫生相关性：我们的目标是基于硫酸软骨素蛋白多糖的强烈抑制特性开发治疗中枢神经系统轴突损伤的新疗法，硫酸软骨素蛋白多糖是一组由反应性胶质瘢痕产生的细胞外基质分子。开发针对这些轴突生长抑制物的新型多肽拮抗剂可能会提高我们治疗成年哺乳动物中枢神经系统轴突损伤的能力。我们希望我们的新治疗策略从体外神经元培养到体内小鼠模型的转换最终将在脊髓损伤和其他中枢神经系统损伤的患者中产生关键策略。

DESCRIPTION (provided by applicant): After CNS axonal injuries, medical treatments to enhance recovery from neurological deficits are extremely limited. Non-permissive environments for axonal growth at least partially contribute to growth failure in the adult CNS. Specifically, several groups of inhibitory molecules strongly suppress axonal extension following CNS lesions, including chondroitin sulfate proteoglycans (CSPGs) generated by glial scars. CSPGs are the principal inhibitory components of glial scars and form a major barrier to regenerating axons. Although several strategies have been reported, digestion of CSPGs with local application of bacterial chondroitinase ABC is the major in vivo approach to surmount growth inhibition of CSPGs after CNS injuries. Important disadvantages, however, preclude the use of this enzyme as a therapeutic option for axonal injury patients, including incomplete removal of inhibitory components from CSPGs, short-period of enzymatic activity at body temperature and inability to cross the blood-brain barrier. In this proposal, we aim to develop novel strategies for treating CNS axonal injury based on inhibition of CSPGs alone or in combination with our previously identified approaches. We hypothesize that peptide antagonists of CSPGs will augment both morphological and functional recovery in a mouse model of CNS injury. Using a bioinformatics approach to define the conserved elements of several CSPGs, we have identified two selective peptide antagonists for CSPGs. Our preliminary studies suggest that these peptides at low nanomolar concentrations principally overcome neurite growth restrictions of CSPGs in neuronal cultures. Systemic application of a CSPG-blocking peptide significantly improves behavioral recovery in CNS axon-injured mice in vivo. In this study, we will characterize the therapeutic potential of these CSPG antagonistic peptides in mouse spinal cord injury (SCI) model. In addition to CSPGs, a number of inhibitory molecules contribute to axonal growth suppression intracellularly mediated via activation of convergent RhoA or glycogen synthase kinase 32 (GSK-32). Recently, we have demonstrated that inactivation of RhoA with ibuprofen or GSK-32 with lithium overcomes growth inhibition of different molecules and significantly promotes axonal growth of descending motor neurons and locomotor recovery in SCI rodents. Thus, we also aim to stimulate a more dramatic axonal regeneration in SCI mice by combining a CSPG-blocking peptide with RhoA-inhibiting ibuprofen or GSK-32-inactivating lithium, two drugs widely used in humans. The use of our novel antagonists for CSPGs, alone or in combination with ibuprofen or lithium, may significantly advance our ability to treat CNS axonal injuries in adult mammals by promoting axonal regeneration and functional recovery. PUBLIC HEALTH RELEVANCE: We aim to develop novel therapies for CNS axonal injuries based on strong inhibitory properties of chondroitin sulfate proteoglycans, a group of extracellular matrix molecules generated by reactive glial scars. Development of novel peptide antagonists for these axonal growth inhibitors may advance our ability to treat CNS axonal injuries in the adult mammals. We hope that the translation of our novel therapeutic strategies from neuronal cultures in vitro to mouse model in vivo will ultimately lead to key strategies in patients with spinal cord injury and other CNS lesions.