运动干预对皮质脊髓束缺失小鼠脊髓局部微环境和神经网络可塑性的影响

The corticospinal tract injury is the leading cause of motor disabilities in central nervous system (CNS) diseases. Prenatal damage of the corticospinal tract commonly happens in the patients with cerebral palsy and exercise training is the main treatment for motor rehabilitation in clinic. Due to lacking available animal models, the mechanisms of exercise roles remain elusive, which also limits other subsidiary intervention to be used. Using Celsr3 conditional knockout techniques, we generated a novel animal model in which the corticospinal tract is genetically absent. These animals maintain the spontaneous plasticity of reorganizing spinal neural network, which plasticity contributes to partial functional compensation, but these mice still have the deficits in maturation of motor-related components and behaviors. Exercise training is confirmed to promote neural plasticity and functional recovery in adult spinal cord injury. Thus, our hypothesis is: Exercise training may affect spinal microenviroment, which enhances plastic changes of spinal neural network and rescues behavioral impairments in the mice without the corticospinal tract. To test this hypothesis, the animals will take exercise training, and we then study microenviroment changes，the morphological maturity of motor-related components, the plastic changes of spinal networks and functional recoveries in the spinal cord. Our studies will illustrate the mechanisms of exercise intervention from the morphology, function evaluation and related molecular signaling, which results will provide the basis for the clinic therapy of exercise intervention in the patients with the corticospinal tract injuries, such as cerebral palsy and spinal cord injury，and help to find new intervention targets.

皮质脊髓束受损是中枢神经系统疾病运动障碍的主要原因。发育期皮质脊髓束受损常见于脑瘫患者，临床最主要的治疗为运动康复。因缺乏有效的动物模型，运动治疗效果的机理缺乏深入了解，也限制其它辅助干预措施的应用。通过Celsr3条件性敲除，我们建立了遗传性皮质脊髓束剥夺小鼠模型，发现其脊髓神经网络具有自发重塑潜能，并实现功能的部分代偿，但其运动相关神经结构、行为学依然存在缺陷。鉴于运动训练在成年脊髓损伤被证实能促进神经可塑性和功能恢复，本项目提出：运动训练可以改善皮质脊髓束缺失小鼠脊髓的局部微环境，促进脊髓神经网络可塑变化并纠正行为学缺陷。我们拟对该小鼠进行跑步训练，研究脊髓局部微环境变化、运动相关神经结构成熟、脊髓神经网络可塑性和功能恢复。研究结果将从形态结构、功能评价和细胞分子信号三方面阐述运动干预的机理，成果为如脑瘫、脊髓损伤等皮质脊髓束受损患者提供治疗的理论依据，也为探索新的干预靶点奠定基础。

发育期皮质脊髓束受损常见于脑瘫患者，临床最主要的治疗为运动康复训练。通过Celsr3条件性敲除，我们建立了遗传性皮质脊髓束剥夺小鼠模型Emx1–Cre;Celsr3f/−。本项目，我们将成年Emx1–Cre;Celsr3f/−小鼠随机分为运动组和非运动组，持续观察了12周的自主跑轮运动对其功能改善的贡献和潜在机制。运动组小鼠经过8周运动后，其拉力大小，爬网格错误率和食丸抓取检测评分均有明显改善。形态学研究进一步支持了这些结果：运动组小鼠神经肌肉接头数量增加了16％，伴有肌肉重量和体积的增加，且其肌电图记录显示其振幅和波宽分别增加48.2%和43.8%。提示运动训练可以明显提高Emx1–Cre;Celsr3f/− 小鼠肌张力水平和精细运动能力。同时，运动组小鼠脊髓节段单胺能神经纤维和红核脊髓投射的终末分支均有显著增加，EDU标记所示的神经增殖细胞数目增多4.8倍，且超过90％的阳性细胞为少突胶质细胞。提示运动训练可以促进Emx1–Cre;Celsr3f/− 小鼠脊髓神经网络可塑性。此外，跑步运动后，全基因组转录水平测序结果提示，差异基因富集于代谢、增殖等生物过程；通路分析显示差异基因主要富集通路与吞噬体以及间隙连接（电突触）相关；蛋白印记实验表明，运动后小鼠脊髓组织突触相关蛋白表达上升，髓鞘碱性蛋白表达升高；Bioplex检测结果显示脊髓局部炎症相关因子水平没有发生明显改变。提示运动可以改善Emx1–Cre;Celsr3f/− 小鼠脊髓局部微环境。总结：自主跑轮运动可以增多单胺能神经纤维和红核脊髓投射的终末分支，促进少突胶质细胞发生，提高突触相关蛋白和髓鞘相关蛋白的表达水平，改善皮质脊髓束缺失小鼠的精细运动功能。这为我们提供了脑瘫临床运动康复的基本知识。