Id2与ARSB联合应用促进脊髓损伤后轴突再生和功能恢复的实验研究

The failure of axonal regeneration following adult spinal cord injury (SCI) has been mainly attributed to the diminished intrinsic regenerative capacity of mature neuron and the extrinsic inhibitory environment. We previously showed that transcription regulator Id2 can stimulate the intrinsic growth ability of adult neuron, however it failed to overcome the inhibition of CSPGs, which is one of the key players in preventing axonal regeneration. In an effort to find strategies to overcome CSPG inhibition, we found arylsulfatase B, an enzyme clinically used for treatment of Mucopolysaccharidosis type VI, can hydrolyze sulfate groups of N-Acetyl-D-galactosamine in the glycosaminoglycan side chains of CSPGs and hence reduce its inhibitory effect on neurite outgrowth. Based on these findings, we hypothesize that: a combinatorial treatment with Id2 to stimulate intrinsic regenerative capacity plus ARSB to block extrinsic CSPG inhibition can synergistically boost the axonal regeneration, circuits remodeling and functional recovery following SCI. In this proposal, we aimed to test this hypothesis both in primary cell cultures and in a mouse dorsal spinal cord hemisection model. We will assess the combination therapy on axonal regeneration, synaptic remodeling as well as functional recovery. We expect enhanced axonal regeneration and functional recovery will be achieved by this combination therapy. This study may provide a novel strategy for the treatment of SCI.

通过刺激轴突再生来重塑代偿性神经环路和实现功能恢复是脊髓损伤修复的重要研究方向。成年中枢神经不能再生的原因主要有神经元本身缺乏再生能力和损伤微环境存在硫酸软骨素蛋白聚糖(CSPGs)等再生抑制分子。我们前期研究表明：转录调控因子Id2可激发成年神经元内在活力，但不足以克服损伤处堆积的CSPG对轴突再生的屏障作用，提示有必要联合消除CSPGs抑制作用的策略。我们体外研究发现：临床上用于治疗粘多糖贮积症的芳基硫酸酯酶B(ARSB)可水解CSPGs上的硫酸酯基团从而消除其对轴突再生的抑制。因此我们提出假说：提高神经元Id2的表达并联合ARSB酶解损伤处的CSPGs可促进脊髓损伤后的轴突再生和功能恢复。本项目旨在细胞实验和脊髓损伤动物模型中明确Id2与ARSB联合应用的效果，包括对轴突再生、突触重建及动物功能恢复情况进行评估；并探讨Id2和ARSB的作用机制。该研究有望为脊髓损伤修复提供新策略。

通过重塑代偿性神经环路来实现功能恢复是脊髓损伤修复的一个重要研究方向。造成脊髓损伤再生失败的原因主要有：1）成年神经元自身缺乏生长能力；2）损伤局部存在硫酸软骨素蛋白聚糖（CSPGs）等抑制分子，不利于轴突再生。因此要促进轴突再生并重建神经环路需要在提高神经元内在活力的同时消除CSPGs等的抑制。前期发现：临床药物芳基硫酸酯酶ARSB可降解CSPGs的硫酸酯基团，而转录调控因子Id2可激发成年神经元内在生长活力。在本项目中，我们利用体外细胞实验及体内脊髓损伤动物模型开展系列研究，主要取得了如下研究进展：1）进一步明确了Id2对神经元轴突生长的促进作用，并发现Id2通过上调Neurogenin2来促进轴突生长这一新机制；2）证明提高星形胶质细胞中ARSB的表达可促进神经元轴突生长，而降低星形胶质ARSB的表达抑制轴突生长；3）单用Id2无法有效克服CSPGs的抑制作用，而联合应用Id2和ARSB可使神经元有效克服CSPGs对轴突生长的抑制作用；4）在胸段脊髓损伤后，过表达Id2可促进脊髓损伤下游腰段脊髓内的轴突代偿性出芽及环路重建；5）在脊髓损伤区域过表达ARSB可促进皮质脊髓束的再生和动物运功功能恢复。我们通过本项目的研究为脊髓损伤轴突再生找到了Neuregenin2这一新的靶点，并证明临床药物ARSB有促进脊髓损伤后轴突再生的效果，具备一定的转化前景。