基于钠离子通道研究东亚钳蝎蝎毒多肽修复脊髓损伤的作用机制

Spinal cord injury (SCI) is a disease with high morbidity and disability rate. whereas the current treatment methods can not meet the clinical needs. Promoting neurite growth is one of the strategies to treat SCI. Buthus martensii Karsch (BmK), one of Traditional Chinese Medicine, has been used to treat hemiplegia, facial paralysis, and other neurological diseases in the clinic. Our previous study indicated that BmK NT3, a peptide obtained from BmK, was able to promote neurite outgrowth subsequent to sodium channel activation in cultured spinal cord neurons. What’s more, BmK NT3 enhanced the motor function score of SCI mice, and improved the pathological status of injured spinal cord. Therefore, this project proposes the hypothesis that BmK NT3 promotes neurite outgrowth and repairs spinal cord injury by activating sodium channels. To testify this propose, electrophysiological technique will be used to demonstrate the agonistic effect of BmK NT3 on sodium channels. Meanwhile, pharmacological and biological methods will be applied to study the mechanism of BmK NT3 promoting neurite outgrowth. Finally, an SCI mice model will be established to demonstrate the in vivo efficacy of BmK NT3 in repairing SCI. This project will preliminarily clarify the regulation effect of sodium channels on neurite outgrowth, and prove the potential of sodium channel as a novel therapeutic target for SCI treatment, as well as reveal the mechanism of BmK NT3 in repairing SCI. In addition, this project will provide theoretical basis for the traditional efficacy of BmK.

脊髓损伤（SCI）是一种高发病率和高致残率的疾病，现有治疗手段不能满足临床需求。促进神经突生长是治疗脊髓损伤的策略之一。东亚钳蝎在临床上用于治疗半身不遂、面瘫等神经系统疾病。课题组前期研究发现东亚钳蝎蝎毒多肽BmK NT3能够激动钠通道进而促进背根神经元神经突生长，还能够提高SCI小鼠的运动功能评分，改善受损脊髓的病理状态。因此，本项目提出“BmK NT3通过激动钠离子通道促进神经突生长从而修复脊髓损伤”的假说。本项目将利用膜片钳技术证明BmK NT3对钠通道的激动作用；同时利用药理学和分子生物学手段研究BmK NT3促进神经突生长的作用机制；最后建立SCI模型证明BmK NT3修复脊髓损伤的体内效能。本项目将初步阐明钠离子通道对神经突生长的调控作用，证明钠离子通道有可能能够成为一个治疗脊髓损伤的新颖作用靶点；揭示BmK NT3修复脊髓损伤的作用机制；并为东亚钳蝎的传统功效提供理论依据。

脊髓损伤（spinal cord injury，SCI）是一种高发病率和高致残率的疾病，现有治疗手段不能满足临床需求。促进神经突生长是治疗脊髓损伤的策略之一。东亚钳蝎在临床上用于治疗半身不遂、面瘫等神经系统疾病。课题组前期研究发现东亚钳蝎蝎毒多肽BmK NT3能够激动钠通道进而促进背根神经元神经突生长，还能够提高SCI小鼠的运动功能评分，改善受损脊髓的病理状态。因此，本研究以东亚钳蝎蝎毒多肽中的活性成份BmK NT3（后根据该多肽功能命名为Buthus martensii Karsch neurite-stimulating peptide，BmK NSP）为研究对象，通过体内、体外实验确证东亚钳蝎蝎毒多肽BmK NT3通过上调脑源性神经营养因子（brain derived neurotrophic factor，BDNF）分泌能够促进脊髓神经元轴突生长，初步阐明蝎毒注射液对脊髓损伤治疗作用的物质基础与新颖机制。研究结果：（1）BmK NT3显著延缓电压门控钠离子通道（voltage-gated sodium channel，VGSC）的失活，从而使通道持续开放，对Nav1.2通道的影响效力最大；（2）使用VGSC抑制剂河豚毒素（Tetrodotoxin，TTX）证明了BmK NT3调节脊髓神经元轴突生长是由VGSC介导的；（3）BmK NT3使脊髓神经元BDNF基因水平表达增加、分泌增多，使用Trk B高效选择性抑制剂可显著抑制BmK NT3诱导的脊髓神经元轴突的生长；（4）为了进一步明确东亚钳蝎蝎毒多肽修复脊髓损伤的物质基础与作用机制，本项目进一步从东亚钳蝎蝎毒中分离得到活性多肽BmK NT1与BmK NSPK，依托于本项目建立的筛选模型与研究方法，发现了东亚钳蝎蝎毒中的活性多肽，并初步阐明了钠离子通道对神经突生长的调控作用，证明钠离子通道有可能成为一个治疗脊髓损伤的新颖作用靶点；揭示BmK NT3修复脊髓损伤的作用机制；本研究为东亚钳蝎的传统功效提供了理论依据。明确了“BmK NT3通过激动钠离子通道释放BDNF，通过BDNF-Trk B途径促进神经突生长从而修复脊髓损伤”的作用机制。