Overcoming glial scar inhibitions on axonal growth

描述（由申请人提供）：CNS轴突损伤后，用于促进神经功能缺损恢复的药物治疗非常有限。轴突生长的非许可环境至少部分导致成年CNS的生长失败。具体而言，几组抑制性分子强烈抑制CNS损伤后的轴突延伸，包括由神经胶质瘢痕产生的硫酸软骨素蛋白聚糖（CSPG）。CSPG是神经胶质瘢痕的主要抑制成分，并形成再生轴突的主要屏障。虽然已经报道了几种策略，但局部应用细菌软骨素酶ABC消化CSPG是克服CNS损伤后CSPG生长抑制的主要体内方法。然而，重要的缺点排除了使用这种酶作为轴突损伤患者的治疗选择，包括从CSPG中不完全去除抑制组分，在体温下酶活性的短时间和不能穿过血脑屏障。在这个建议中，我们的目标是开发新的策略，用于治疗中枢神经系统轴突损伤的基础上抑制CSPGs单独或与我们以前确定的方法相结合。我们推测，肽拮抗剂的CSPGs将增加形态和功能的恢复，在小鼠模型的中枢神经系统损伤。使用生物信息学方法来定义几个CSPGs的保守元件，我们已经确定了两个选择性肽拮抗剂的CSPGs。我们的初步研究表明，这些肽在低纳摩尔浓度主要克服神经元培养物中CSPGs的轴突生长限制。全身应用CSPG阻断肽显著改善CNS轴突损伤小鼠体内的行为恢复。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。除了CSPG之外，许多抑制性分子有助于通过会聚RhoA或糖原合成酶激酶32（GSK-32）的激活在细胞内介导的轴突生长抑制。最近，我们已经证明，用布洛芬或GSK-32与锂灭活RhoA克服了不同分子的生长抑制，并显着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，我们还旨在通过将CSPG阻断肽与RhoA抑制布洛芬或GSK-32失活锂（两种广泛用于人类的药物）组合，刺激SCI小鼠中更显著的轴突再生。单独或与布洛芬或锂组合使用我们的CSPG的新型拮抗剂，可以通过促进轴突再生和功能恢复来显著提高我们治疗成年哺乳动物CNS轴突损伤的能力。 公共卫生相关性：我们的目标是开发新的治疗中枢神经系统轴突损伤的硫酸软骨素蛋白聚糖，一组细胞外基质分子反应性胶质瘢痕产生的强抑制特性的基础上。开发这些轴突生长抑制剂的新型肽拮抗剂可能会提高我们治疗成年哺乳动物CNS轴突损伤的能力。我们希望我们的新的治疗策略从体外神经元培养到体内小鼠模型的翻译将最终导致脊髓损伤和其他CNS病变患者的关键策略。

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.